Anatomy

TMNT Anatomy: The Weirdest Things About the Ninja Turtles’ Bodies – CBR – Comic Book Resources

The Teenage Mutant Ninja Turtleswere created by Kevin Eastman and Peter Laird and first appeared in Mirage Comics in 1984. Since then they've appeared in television shows, movies, video games and comic books. TMNT has been entertaining people for decades and every generation has looked different, but Leonardo, Donatello, Michelangelo and Raphael have always been there with their humanoid turtle anatomy, kicking butt.

If there were any doubts regarding the turtle's superhero nature, those have squashed thanks to recent team-ups with Batman and the Power Rangers along the way. TheTurtles' super abilities aren't put on blast like many other popular heroes, but that doesn't mean they don't exist.

Related:TMNT #101 Is the Ideal Jumping-On Point Heres Why

The TMNT were created by a mysterious radioactive ooze that mutated them into humanoid turtle creatures. The mutations culminatedin heavily armored features like their shells and cranial plates. Their hands became tridactyl forelimb with opposable thumbs that allow them to maneuver their weapons and their feet are similar, but only have two digits. The Turtles's internal organs resemble human anatomy,including a chambered heart structure and similar intestinal system. They also possess the emotional depth of a human and many human interests and tastes -- like pizza, for one.

The Turtles have always possessed meta-human abilities like enhanced strength, speed, agility and reflexes. They're also resistant to cold. Splinter's ninjutsu training allowed the turtles to hone and strengthen their abilities to incredible levels. Leonardo even took his ninjutsu training to otherworldly heights by obtaining the ability to heal himself and others from fatal wounds. This healing ability goes beyond theTurtles' amphibious healing properties. The height of this power allowed Leo to cure himself of a so-called incurable venom when he was fighting Shredder's estranged daughter.

Related:Two Teenage Mutant Ninja Turtles Villains Just Became Much Bigger Threats

While their muscle and skeletal structures combine many elements from turtles and humans,the Teenage Mutant Ninja Turtles' shellsare incredibly similar to real turtle shells. The plastron and the carapace combine to cover the Turtles' front and back to cover all their vital organs. Even the simpler animations show the shells divided into smaller subsections, whichare points where the individual bones fuse and form into a shell.

Their shells take the place of their abdominal muscles and collar bones and relay a lot of synchronicity between their human and turtle anatomy. Their shells are bulletproof and can withstand enormous kinetic force, making them incredibly resistant in battle -- which allows them to more easily defend against multiple opponents at once. The shells also provide situational offensive capabilities, like when the turtles hurl themselves like cannonballs at their foes.

Related: Teenage Mutant Ninja Turtles: IDW's Urban Legends Saved TMNT's Lost Era

Jennika is the fifth member of the TMNT and is objectively the most unique turtle of the bunch. Not only was she human at one time, but she also worked for Shredder was one of Splinter and theTurtles' most deadly foes. After she was gravely wounded, the Turtles decided to save her life with a blood transfusion. Leo was Jennika's blood donor and his mutated blood transformed Jennika into a turtleoid, whereas the original turtles were turned into humanoids. Jennika's Turtle form is far more slender than her male counterparts and she dawns a bright yellow eye mask to complete her look.

Jennika's main weapon choice is a set of claws, keeping an element of her Foot Clan past with her during her new life as a hero in a half shell. It's Jennika's mental fortitude that sets her apart. Not only does she have an entire lifetime of human memories, she's dealing with all of the mental hurdles of a mid-life species change. Jennika brings a lot more weight and human nuance to her character, which makes her a completely new breed ofMutant Ninja Turtle.

Related:TMNT: Jennika #1 Offers a Punk-Rock Look at the Newest Hero in a Half Shell

The turtles have gone through some changes over the years. While they generally remain the same, there have been some radical exceptions to the rule. One time, Leo lost his hand in a battle with a mutated Komodo Dragon named King Komode and in the same comics series, Raphael's face became permanently scarred. These incidents are no longer considered canon, but theyprovide an idea of theTurtles' healing limitations and the extent of their pain tolerance.

Donny and Raph have gone through the weirdest changes of them all, like the one time they turned into vampires during a collision with Universal Studio's classic monsters-- including Dracula. Donny also becomes a cyborg at one time when his body is damaged from a nearly fatal fall out of a helicopter. Donny merged with one of the cyborgs he and his brothers were fighting to save his life and the CPU altered his brain to make him more aggressive and feel less remorse. His extra mechanical features included blasters and improved armor.

The Teenage Mutant Ninja Turtles may have started as turtles mutated into humanoids by toxic ooze, but they have come a long way since and are now genuine heroes with incredible power.

Keep Reading:How the Ninja Turtles Come to Terms with [SPOILER's] Death

Dragon Ball Super Reveals the TRUTH About Goku's Ultra Instinct Abilities

Jonathan Olsen-Koziol has been training for this his entire life. Since he was a young child he's been consuming and studying comic books, movies, video games, heroes, villains, story arcs, and pop-culture. Now he gets to relay a lifetime of passion and knowledge to the people! If his writings find you; hopefully you leave entertained. Jon graduated from Central Washington University with a Bachelor's in journalism and concurrently works as a head editor and digital content specialist for RespectMyRegion.com.

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TMNT Anatomy: The Weirdest Things About the Ninja Turtles' Bodies - CBR - Comic Book Resources

Anatomy

The anatomy of failure: How and why the emergency alert system was not activated when a mass murderer was roaming around Nova Scotia – Halifax…

Lillian Hyslop. Photo: Facebook.

Wentworth Valley resident Heather Matthews believes her walking companion might still be alive if the RCMP or Nova Scotias Emergency Measures Organization (EMO) had issued a special alert early Sunday morning to stay inside. Instead, she says Wentworth resident Lillian Hyslop a 60 year old, community-minded person, and parent was gunned down during her daily walk by a killer who had already taken multiple victims and was the subject of a 10-hour manhunt.

If there is ever hopefully never a next time let people know, urges Matthews. Theres so many different reasons why we should be given an alert that could save lives. They give us the Amber alert, we get the COVID-19 alert twice over the Easter weekend?

Heather Matthews lives above the former Highway 104, known locally as The Four, that runs past the Wentworth Ski Hill north toward Oxford. She and her husband David were walking Sunday morning along a forested path parallel to the road when they heard what they described as a single gunshot. Heather estimates the time of the shooting at 9:20-9:30am because she had checked the clock before leaving home.

When the couple returned home, they had a phone call from a neighbour warning them to lock their doors and not go outside because there were reports a gunman was in the area. Heather Matthews immediately called Lillian Hyslops mother-in-law to tell her, but it was too late. Lillian had already left for her daily walk. She died near the entrance to the Wentworth Provincial Park.

I really believe if there had have been an alert sent out, some of these random people that were shot would not have been shot, Matthews told the Halifax Examiner. The people he had targeted, thats one thing. But these people he got just because they were there?

The women [employed by the VON] like Kristen Beaton and Heather OBrien who were out driving. Perhaps if they had been given warning, Matthews voice trailed off.

There are also questions in the community about whether Tom Bagley, 70, would have died if an alert had been issued over the cellphone network or the radio. Bagley was out for a walk when he saw a house burning on Hunter Road. Two people inside, Corrections Canada managers Sean McLeod and Alanna Jenkins, perished in the blaze. The Wentworth Fire Department was notified about the fire, but firefighters were instructed not to respond because the situation was too dangerous. Bagley didnt know that. The retired firefighter with the Canadian Armed Forces went to help and was shot and killed by the gunman.

At 9pm on Saturday night, the RCMP communications people were evidently in a good mood. They posted a fun tweet, which in hindsight is eerily sad:

Its Saturday and our #9PMROUTINE is complete! That means we get to relax for the evening and sleep in tomorrow. Are you done your #lockup? When you are, share this post! pic.twitter.com/WBfYWsUDNn

RCMP, Nova Scotia (@RCMPNS) April 19, 2020

An hour and a half later, at 10:26pm, RCMP officers responded to a possible shooting in Portapique, said RCMP Chief Superintendent Chris Leather at a press conference Tuesday afternoon.

At about 11:08 pm, a first responder at Portapique reported that:

So theres a structure fire. Theres a person down there with a gun. Theyre still looking for him. The patient we have got shot by him. He was just down there observing the fire, checking out the fire, so there could be other patients around the fire that could be gone already, but were not sure. Police are stationed at the end of the road there on the 2, not letting anybody down any further but its very vague whats going on down there but there is for sure multiple patients down there.

So police knew there were multiple people shot and the gunman was on the loose, but at about 11:30pm, the RCMP tweeted only that there was a firearms complaint in the Portapique area.

#RCMPNS is responding to a firearms complaint in the #Portapique area. (Portapique Beach Rd, Bay Shore Rd and Five Houses Rd.) The public is asked to avoid the area and stay in their homes with doors locked at this time.

RCMP, Nova Scotia (@RCMPNS) April 19, 2020

Leather said that the search for the gunman continued into the evening and early morning hours on Sunday. In response to new information indicating the suspect was not in the secure perimeter [around Portapique], at 8:02am on Sunday, the RCMP began providing real-time information on its Nova Scotia RCMP Twitter account.

But the 8:02am tweet implied only that the shooter was in the Portapique area:

#RCMPNS remains on scene in #Portapique. This is an activeshooter situation. Residents in the area, stay inside your homes & lock yourdoors. Call 911 if there is anyone on your property. You may not see the policebut we are there with you #Portapique.

RCMP, Nova Scotia (@RCMPNS) April 19, 2020

The next tweet from the RCMP came at 8:54 AM, and it carried a photo of the suspect along with this message. The only hash-tagged location in the message was Portapique.

Around this time, there was the house fire on Hunter Road in Wentworth, over 40 kilometres from Portapique, where the two corrections officers Alanna Jenkins and Sean McLeod lived. Only later, on Monday, would the RCMP add their names to the list of the perpetrators victims, along with that of Tom Bagley, the 70-year-old neighbour and retired firefighter who went to the burning house to see if he could help, only to be shot and killed. And Lillian Hyslop was killed while walking along The Four.

At that point, there had been no mention in RCMP tweets that the killer had left Portapique, or of any danger in Wentworth, and yet the RCMP told firefighters not to attend the scene because of the danger of the gunman.

The next tweet came at 10:04am:

#RCMPNS is advising people to avoid Hwy 4 near Hidden Hilltop Campground in #Glenholme. Gabriel Wortman is in the area. Please stay inside your homes and lock your doors. #Portapique

RCMP, Nova Scotia (@RCMPNS) April 19, 2020

The very first reply to that tweet came from MaritimeGirL who wrote:

This should be announced on the emergency alert system. People out there driving may have no clue.

At 10:13am Sunday first responders reported two cars in MVA (motor vehicle accident) with possible entrapment (people trapped in the vehicles) at 1760 Plains Road in Debert:

At 10:15am, said Leather, the Emergency Management Office (EMO) contacted the RCMP to offer the use of the public emergency alerting system. We were in the process of preparing an alert when the gunman was shot and killed by the RCMP.

The problem with that explanation is that an hour and 25 minutes elapsed between 10:15 and the time the Twitter account announced that the gunman was in custody (curiously, the account did not say the gunman was killed).

During that period, nine more tweets were issued.

Two connected tweets at 10:17am and 10:21am:

Gabriel Wortman is currently in the #CentralOnslow #Debert area in a vehicle that may resemble what appears to be an RCMP vehicle & may be wearing what appears to be an RCMP uniform. Please stay inside and avoid the area. #RCMPNS

RCMP, Nova Scotia (@RCMPNS) April 19, 2020

Those tweets elicited an immediate tweeted reply from Joanne Roberge, who tagged Premier Stephen McNeil:

If there was ever a time to send out an emergency sound off alert to everyones phone it is now! You sent it out on Easter to have people stay home this is just as serious.

Another two at 10:39am:

Thank you for your understanding as we work to provide the most updated information while addressing public and officer safety. 2/2 #Portapique #CentralOnslow #Debert #Glenholme #Colchester

RCMP, Nova Scotia (@RCMPNS) April 19, 2020

And another one at 11:04am:

Gabriel Wortman, suspect in active shooter investigation, last seen travelling southbound on Hwy #102 from #Brookfield area in what appears to be RCMP vehicle & may be wearing RCMP uniform. Suspect's car is 28B11, behind rear passenger window. If you see 28B11 call 911.

RCMP, Nova Scotia (@RCMPNS) April 19, 2020

Once again there was an immediate reply from a follower Levi Marshall asking for an alert:

Can we get an emergency alert please! Not everyone checks their phones after waking up?

Yet another tweet at 11:06am:

Gabriel Wortman, suspect in active shooter investigation, now believed to be driving small silver Chevrolet SUV. Travelling southbound on Hwy #102 from #Brookfield area If seen, call 911.

RCMP, Nova Scotia (@RCMPNS) April 19, 2020

At that time, Joey Webber was on a family errand towards Shubenacadie, Halifax councillor Steve Streatch told the Halifax Examiner. [Joeys] father told me this morning that [Joey] came right up around the ramp where the police cars were on fire there. He just left the service station to come up around, and he met the face of evil.

Joey was killed. The gunman took the silver Chevy Tracker Joey had been driving.

The RCMP tweeted out at 11:24am:

Confirmed suspect vehicle is silver Chevy Tracker. Last seen #Milford. If seen, call 911.

RCMP, Nova Scotia (@RCMPNS) April 19, 2020

Then at 11:35am:

To clarify, the suspect in our active shooter investigation, Gabriel Wortman, is NOT employed by the RCMP but he may be wearing an RCMP uniform. He is considered armed and dangerous. If you see him, do NOT approach and call 911 immediately.

RCMP, Nova Scotia (@RCMPNS) April 19, 2020

And finally, at 11:40am:

Gabriel Wortman, suspect in active shooter investigation, is now in custody. More information will be released when available. Thank you for your cooperation and support. #Colchester

RCMP, Nova Scotia (@RCMPNS) April 19, 2020

Oddly, the final tweet did not mention that the gunman was killed.

At todays daily COVID-19 briefing, Canadian Press reporter Keith Doucette had the following exchange with Premier Stephen McNeil:

Doucette: The US Consulate in Halifax issued an email alert to their citizens on Sunday about an active shooter based on the Nova Scotia RCMPs Twitter feed. Why was the province waiting for official word from the Mounties before issuing an emergency alert?

McNeil: Well, thats the protocol in place when it comes to EMO. As you know, the emergency alert, the lead agency is the one who has to put the message together. We would not go by whats happening by Twitter. We would need the lead agency to actually craft the message so that we could put that out and no message was received even though EMO had reached out a number of times throughout the morning to the RCMP.

Doucette: Why did you have EMO staff in anyway? It appears that you were anticipating that something was needed and then nothing happened. Is that a question for you, that youre wondering about?

McNeil: That command centre wouldve been going around COVID, so we stood that on early on, EMO put that up. But through a number of things that we were seeing on, or the EMO staff was seeing on Twitter, they called in their support staff around the alert message team and then began to reach out to the RCMP, who would have to craft the message or work with our team to craft that message and no message was received.

This morning, the Emergency Management Office confirmed to the Halifax Examiner that the Provincial Coordination Centre (PCC), from which emergency alerts can be issued, has been staffed seven days a week (but not nights) since the COVID-19 State of Emergency was declared on March 22.

The Halifax Examiner was not provided the opportunity to ask a question at the COVID-19 briefing. But we wanted to follow up the exchange between Doucette and McNeil by asking: What exactly is the emergency alert system for, if not for making the quick decision to alert the public to imminent danger?

McNeil seemed to be describing a complex alert system, such that the PCC first has to be staffed, and then get an ask from whatever the lead agency is in the case of an emergency. Then, the lead agency, perhaps in cooperation with the EMO, has to craft an alert, and only then can the alert be activated.

Is this bureaucratic system overly complex? What if the emergency happens at night, or otherwise when the PCC isnt staffed? More to the point, what happens if the emergency the public is to be alerted about has a timeframe shorter than the hour and 25 minutes it took between the EMO asking the RCMP to request an alert and the RCMP crafting it? What if there are only moments to spare, as in the case of a tsunami, or tornado, or missile attack or, as it turns out, a mass murderer on the loose?

Can the alert system serve to alert anyone to a real impending emergency?

Terry Canning thinks the system can work just fine; it just didnt this time. Canning was the emergency communications coordinator for the province for 15 years. Before that he was a specialist inmulti-agency trunked mobile radio systems. He is now a radio communications consultant.

We contacted Canning to ask about the lack of an emergency alert during Sundays murder spree.

Somebody within the Royal Constabulary fucked up big time, said Canning in an interview with the Examiner Wednesday evening. And relying on fucking Twitter of all things.

Canning said he has no immediate knowledge of events over the weekend, but in my experience, and Ive been in the emergency response field for 35 years, you would appoint a public information officer pretty quickly in a situation like that. And one of the first responsibilities of a PIO is to distribute information. Obviously, the most efficient way to do that was through the provincial public alert system, without a doubt.

Canning said that even when the PCC is closed, an EMO staffer is on call 24 hours a day, and they have to be within landline or cellphone range during their shift. They also carry radios.

He couldnt say for sure if the off-hours staffer can issue an alert without travelling to the PCC, but he thinks it likely.

Id be very surprised if they dont have some means through the internet, obviously, to get on line and issue the alert from wherever they are, but thats outside my scope of knowledge I cant say that categorically, but Id be very surprised if one of the duty officers couldnt be able to do that wherever they happen to be, so long as they have internet service.

The Examiner has requested a copy of the procedures and policies for issuing an emergency alert, but as of publication, the EMO has not provided it.

The Halifax Examiner is an advertising-free, subscriber-supported news site. Your subscription makes this work possible; please subscribe.

Some people have asked that we additionally allow for one-time donations from readers, so weve created that opportunity, via the PayPal button below. We also accept e-transfers, cheques, and donations with your credit card; please contact iris at halifaxexaminer dot ca for details.

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The anatomy of failure: How and why the emergency alert system was not activated when a mass murderer was roaming around Nova Scotia - Halifax...

Anatomy

Okay, Theres a Greys Anatomy Theory Going Around Saying Jackson Might Die – Cosmopolitan

Gilles MingassonGetty Images

Its been a few weeks since Greys Anatomy season 16 ended, but fans are desperately trying to figure out what would have happened if the season got the last few episodes it was supposed to. One wild theory going around involves Jackson Avery, and fans on Reddit think that if the season had gone on as planned, he actually would have died.

Greys ended its season on episode 21. It was supposed to have 25 episodes total. Obviously, the writers have been weaving the Greys and Station 19 storylines together lately, so the new Station 19 episodes hold some clues for what would have happened in those last four episodes of Greys. The synopsis for an upcoming Station 19 episode says, Ben, Jackson and Emmett are put in a precarious and dangerous position with a potential victim.

Doesnt seem like anything that would cause alarm, right? But someone on Reddit wrote this: According to press release for episode 15 for Station 19, Jackson along with Ben and the newbie are put in a dangerous position. I think thats a given that Jackson would have been the one to die.

Okay, Im not sure that its a given, and I dont totally get the leap between Jackson being in a dangerous position and him straight-up dying (unless the internet knows something I dont), but other people on the Reddit thread agreed. That theory makes a lot of sense, one commenter wrote. Also, his character pretty much peaked and died when Sarah Drew left the show.

And even if youre not totally sold on the Station 19 connection, people were saying Jackson was toast before that episode info dropped. On a different Reddit thread totally unrelated to that, fans wrote they think Jackson will be the next to go. So, um, I guess if you see Jackson die early in the next season of Greys, you heard it here first.

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Okay, Theres a Greys Anatomy Theory Going Around Saying Jackson Might Die - Cosmopolitan

Anatomy

Grey’s Anatomy boss reveals two storylines they couldn’t finish – digitalspy.com

Grey's Anatomy season 16 spoilers follow.

Grey's Anatomy showrunner Krista Vernoff has revealed two storylines for season 16 that the show was forced to cut due to the season ending early.

The 21st episode ended up as the season's makeshift finale, but there were originally four more episodes planned.

Speaking to Deadline, Vernoff described two plots that viewers would have seen happen: a storyline that would have explored why Teddy Altman cheated on Owen Hunt, and a storyline involving Andrew DeLuca that would have revisited the human trafficking case.

"We had an episode coming up where we were able to better articulate and better understand what's prompting Teddy's behaviour, and we don't get to air it," the showrunner said.

"Who knows, maybe it's going to change between now and when we'll actually shoot it for season 17, but I feel for Kim Raver.

"The amount of standing is high, and we've left her in a strange place. It's compelling... Why, when she was finally getting her happy ending, did she sabotage [the relationship]? I think it's [a] super-rich area, personally."

Related: Grey's Anatomy boss confirms "reset" ahead of next season after season 16 forced to end early

And speaking about DeLuca's incomplete storyline, Vernoff explained: "The other story that I was really disappointed that we couldn't complete... we did a story where there was a victim of human trafficking like two episodes ago, and DeLuca recognised it but he was in such a mentally compromised, manic state that nobody listened to him and the girl left.

"We had an episode where she comes back, and I am really sad that we can't air that episode this season because it felt important to offer that kind of hope to people who are living that experience.

"I may still complete that story next season."

Recent reports have also suggested that a major character death was planned for the original season finale. Vernoff didn't confirm or deny this, but did say that fans can watch the remaining episodes of Station 19's third season to find clues.

"Ironically, when fans watch episode 15 and 16 of Station 19, they will probably have a feeling of some of what we had planned for [the original Grey's finale]," she explained to TVLine.

One long-running storyline that was thankfully resolved in time was Richard Webber's mysterious illness, which turned out to be cobalt poisoning from a leaky hip replacement.

Grey's Anatomy airs on Thursdays on ABC in the US. It airs on Sky Witness in the UK with selected episodes also available on NOW TV.

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Grey's Anatomy boss reveals two storylines they couldn't finish - digitalspy.com

Anatomy

Greys Anatomy season 17: Is Richard Webber still in danger? Showrunner confirms fate – Express.co.uk

Greys Anatomy was brought to a premature close after its 16th season finished four episodes early. However, in the finale it seemed Richard Webber (James Pickens Jr) was officially on the mend - heres what showrunner Krista Vernoff had to say about this.

Fans of Greys Anatomy were left in turmoil when Richard began experiencing some worrying symptoms.

This all happened when he hallucinated seeing his wife at a conference he was speaking at.

Then he blacked out and collapsed, leading viewers to believe he was in serious danger.

However, Meredith Grey (Ellen Pompeo) was tasked with finding exactly what was wrong with him.

READ MORE:Grey's Anatomy: Is this the last season of Grey's Anatomy?

This was thankfully resolved in episode 21 when she worked out his symptoms were caused by cobalt poisoning from his hip replacement.

Atticus "Link" Lincoln (Chris Carmack) operated on him and seemed to fix the problem but viewers never saw him recovered and well before the season came to an end.

This, therefore, left some concerned he might not be out of the woods just yet.

Thankfully, showrunner Vernoff has now confirmed what happened next to Richard.

Asked by Deadline if this meant he was now safe, she said: "Yes and I thought that that was one of the most amazing things about this diagnosis.

[It] was that the cobalt poisoning thing is real and it really can cause all of those symptoms.

It can cause dementia-like symptoms. It can cause Parkinson's-like symptoms tremors, hallucinations and the amazing thing about it is that once you diagnose it, and you get the leaky hip out of your body, you can recover, totally.

That felt, just as the storyteller, an amazing thing because it let us give Jim Pickens this really rich, rich material, without us having to permanently disable his character.

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Grey's Anatomy season 16: Was Dr Alex Karev going to be killed off?Greys Anatomy season 17 release date: Will there be another series?Grey's Anatomy season 17: Will Jackson Avery leave for Station 19?

I think that that was an amazing ride for the fans, because the outcry of [us] showing symptoms that don't feel like they have cures.

She went on to explain why this made his storyline so gripping for viewers of the show.

Vernoff added to the publication: "There's not really a cure for Parkinson's. There's not a cure for Alzheimer's.

"So I know everyone was in a panic, and so, we got to tell this really satisfying story, and we got to let people know that sometimes, there's another diagnosis for those symptoms.

Which we found fascinating, as a group of writers who write medicine, that it's so rare to see something this satisfying.

Fans will see Richard back to his old self in the upcoming 17th season of the show.

The new outing has officially been confirmed but it is not clear when it will be released yet.

The coronavirus COVID-19 outbreak could see the series delayed from its usual September slot.

Greys Anatomy is available to catch-up on ABC and Sky

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Greys Anatomy season 17: Is Richard Webber still in danger? Showrunner confirms fate - Express.co.uk

Cell Biology

Meis1 protein plays key role in stopping cell division in the heart – News-Medical.Net

UT Southwestern Medical Center scientists have discovered a protein that works with others during development to put the brakes on cell division in the heart, they report today in Nature.

The findings could eventually be used to reverse this developmental block and help heart cells regenerate, offering a whole new way to treat a variety of conditions in which heart muscle becomes damaged, including heart failure caused by viruses, toxins, high blood pressure, or heart attacks.

Current pharmaceutical treatments for heart failure including ACE inhibitors and beta blockers center on trying to stop a vicious cycle of heart muscle loss as strain further damages remaining heart muscle, causing more cells to die, explains UT Southwestern physician-researcher Hesham A. Sadek, M.D., Ph.D., a professor of internal medicine molecular biology, and biophysics. There are no existing treatments to rebuild heart muscle.

Nine years ago, Sadek and his colleagues discovered that mouse hearts can regenerate if they're damaged in the first few days of life, spurred by the division of cardiomyocytes, the cells responsible for a heart's contractile force.

However, this capacity is completely lost by 7 days old, an abrupt turning point in which division of these cells dramatically slows and the cells themselves enlarge. The reasons why these cells gradually slow and stop dividing has been unclear.

Sadek and his team discovered in 2013 that a protein called Meis1, which falls into a category known as transcription factors that regulate the activity of genes, plays a key role in stopping heart cell division.

However, he explains, although deleting this gene in mice extends the window of heart cell division, this effect is transient heart cells missing this gene eventually slow and stop their multiplication.

Consequently, the researchers wondered whether there were redundant mechanisms in place that stop heart cell division even when Meis1 is absent.

Toward that end, they looked to see what other transcription factors might track activity with Meis1 in heart cells as they rapidly divide and then slow to a halt in the days after birth. They quickly discovered one called Hoxb13 that fit the bill.

Other proteins in the Hox family, Sadek notes, have been shown to act as chaperones to Meis1 in other types of cells, ferrying Meis1 into the cell nucleus.

To better understand Hoxb13's role in heart cells, the researchers genetically engineered mice in which the gene that codes for Hoxb13 was deleted.

These mice behaved much like those in which just the gene for Meis1 was deleted the window for heart cell rapid division was increased but still closed within a few weeks.

When the researchers shut off Hoxb13 in adult mouse hearts, their cell division had a brief resurgence, enough to prevent progressive deterioration after an induced heart attack but not enough to promote significant recovery.

However, when the researchers deleted both the genes for Meis1 and Hoxb13, heart cells in these mice appeared to revert to an earlier stage in development, both decreasing in size and multiplying more.

After an induced heart attack, these mice had a rapid improvement in the amount of blood each beat could expel from the heart. Their heart function had almost returned to normal.

With clear evidence that Meis1 and Hoxb13 work together to stop heart cell division in the days after birth, Sadek and his colleagues looked for what might in turn regulate these proteins.

Their experiments suggest that the answer is calcineurin, a protein that's responsible for regulating the activity of other proteins by removing their phosphate groups.

Because calcineurin plays a key role in a variety of diseases and other medical conditions, such as rheumatic arthritis, schizophrenia, diabetes, and organ transplant, several drugs already exist on the market that target this protein.

Conceivably, says Sadek, other drugs could be developed to directly target Meis1 and Hoxb13. Researchers may eventually be able to develop strategies to restart heart cell division through a single drug or combinations that target any part of this regulatory pathway, he adds.

By building up the story of the fundamental mechanisms of heart cell division and what blocks it. we are now significantly closer to being able to harness these pathways to save lives."

Hesham A. Sadek, M.D., Ph.D., Physician-Researcher and Professor of Internal Medicine Molecular Biology, and Biophysics, UT Southwestern Medical Center

Sadek also holds the J. Fred Schoellkopf, Jr. Chair in Cardiology.

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Meis1 protein plays key role in stopping cell division in the heart - News-Medical.Net

Cell Biology

[OPINION] We need to professionalize biology in the Philippines – Rappler

The first 4 months of the year 2020 saw raging forest fires in Australia, the eruption of a volcano (Taal Volcano) located in the middle of a caldera lake (Lake Taal), a locust infestation in parts of Africa, the COVID-19 pandemic which traces its origins to bat and/or pangolin consumption in China, and turquoise-blue waters returning to Manila Bay during the first few weeks of the "enhanced community quarantine." These of course, came at the heels of a water crisis in Metro Manila, the declaration of Sardinella tawilis as an endangered species last year, and the continued negative impacts of climate change.

All these scenarios have brought challenges that have affected all our lives. Social media has increased public interest in these phenomena but have likewise led to many misinterpretations and fake news. One thing shared by all these events is that they are best understood by persons who have degrees and training in biology. They would be among the most knowledgeable members of the population who would be able to understand, explain, offer advice, and have the skills to help study, analyze, and even mitigate the negative impacts of such catastrophic events. Unfortunately, the lack of recognition of biology as a valid career option prevents biology graduates from being appreciated and recognized for their contributions. (READ: Recipe for disaster? U.P. scientists slam release of invasive 'anti-dengue' species)

Studying biology is so much more than the usual high school biology activities such as dissecting frogs or identifying the parts of a flower. Biology is an extremely multi-faceted discipline, from the molecular level all the way to the entire biosphere. The recommended CHED curriculum in biology (CHED CMO 49 s. 2017) provides opportunities to learn basic and applied concepts in taxonomy and systematics, cell and molecular biology, evolutionary biology, genetics, developmental biology, microbiology, and physiology. These are then further strengthened by specialization courses such as bioinformatics, industrial biotechnology, marine biology, conservation biology, medical microbiology, histology and histopathology, parasitology, and immunology, among others. These options largely depend on the chosen specialization track, or major, of the student. A B.Sc. Biology graduate would therefore be knowledgeable in the use of microscopic techniques, DNA isolation, and analysis (which is extremely important in determining positive COVID-19 cases using a PCR machine), microbiological assays, testing on animal models, mapping, statistical analyses, environmental monitoring, and identification of organisms to species level. (READ: Top PH biologist: Tech, data help us understand species better)

Unfortunately, an undergraduate degree in biology, which is offered by 202 Higher Educational Institutions (HEI) in the Philippines (Raymundo et al., 2017) is widely perceived as a dead-end degree. Many students (and even their parents) usually dread career options in biology if in case one decides not to pursue a medical career. Oftentimes, graduates of B.Sc. Biology who do not proceed to medical school end up underemployed unable to practice whatever tangible knowledge or skills they obtained from their undergraduate studies.

It is not surprising to find B.Sc. Biology graduates engaged in entry-level careers in Business Process Outsourcing (i.e. call centers), sales, real estate, and marketing. Others resort to taking a second baccalaureate degree which offers better career prospects, especially abroad, such as nursing or physical therapy. This is because earning a decent living as a biology graduate means taking additional graduate degrees yet eventually competing for a limited number of available job opportunities. Unfortunately, this scenario wastes 4 years of hard toil, spent studying the rigors of biology courses, as well as the research training they obtained while working on their undergraduate thesis. (READ: VIRAL: UP student studying biology while in rally)

I would like to attribute the anxiety of many students over potential career prospects in biology to factors such as 1) the lack of recognition of biology as a valid profession in the Philippines; 2) the competition with, and preference for, graduates of other degrees who have government licensure examinations (medical technologists, chemists, fisheries graduates, agriculturists, foresters, etc.); 3) the low regard for research, innovation, and development among Philippine-based industries and corporations; and 4) the limited job opportunities in academia given the research culture in many higher educational institutions.

In this article, I hope to offer suggestions to address the first two of the 4 factors listed above. The last two factors, though equally important, are not unique to biology and covers other science, technology, engineering, and mathematics (STEM) disciplines. Furthermore, the Department of Science and Technology (DOST) has been continuously coming up with initiatives that have helped ease this problem, such as 1) providing scholarship opportunities for graduate degrees in STEM (DOST-SEI, 2020); 2) raising compensation packages for research staff (DOST, 2019); and 3) strengthening the Scientific Career System for scientists in public service (NAST, 2020). Though far from perfect, this shows how the DOST has been striving to champion the plight of those engaged in S&T careers.

At present, initiatives to professionalize a discipline in biological sciences is limited to microbiology. The Philippine Academy of Microbiology (PAM) offers a certification examination for Registered Microbiologists. The PAM aims to take this to the next level, by soliciting support for a Microbiology Bill in the Philippine legislature (Chipeco, 2019). Once approved, this would transform the exam into a Professional Regulations Commission (PRC) board examination, thus giving microbiologists proper government recognition. This may be used as a benchmark for professionalizing biologists.

Since biologists are needed in many other disciplines apart from microbiology-related professions, the entire discipline needs to undergo proper certification as well. This will enable biologists to have a distinct identity compared to other related professions, which have the unique advantage of filling in vacant positions simply because they have professional board examinations even though these tasks may be best performed by a biologist. To give one possible scenario, a job opening intended for a molecular biologist may be easily filled up by a licensed medical technologist because biology graduates, even though they are obviously qualified to perform the tasks of a molecular biologist, do not hold a government-issued professional license.

However, given the limited job opportunities and the need for further specializations in graduate school, any attempt to come up with a professional certification examination for biologists must specify that a masters degree in biology be the minimum requirement to qualify for the exam, following the example of the Psychology Law (R.A. 10029) for Registered Psychologists. This would ensure that the candidate has taken more than the basic course requirements specified in the undergraduate curriculum and has been exposed to a certain field of specialization in graduate school which makes them more prepared to engage in a highly specialized career. Given how graduate students are encouraged to publish in valid, peer-reviewed journals as part of their graduate training, requiring the completion of a masters degree prior to taking the certification examination ensures that anyone who qualifies as a professional biologist has undergone adequate research training, which is among the strengths of anyone who has pursued advanced degrees in biology.

Another option is to empower biology associations or societies involved in different disciplines to maintain a certain level of quality among their members by providing professional certification programs in various subspecialties or disciplines. This is not without precedent. In Canada, organizations such as the Alberta Society of Professional Biologists (ASPB) and the Association of Professional Biology (APB) in British Columbia regulates and ensures the qualifications of those who wish to practice biology as a profession in their respective federal states. Professional certifications also exist for highly specialized disciplines such as wildlife biology and fisheries biology.

In the Philippines, this means that for us to jumpstart any attempt to professionalize biology, the currently existing professional societies in various fields of biology must provide professional certificate programs in their respective disciplines. They should also come together and form an umbrella organization or academy that would spearhead reaching out to the national government and employers to recognize such professional certifications issued by their respective societies in choosing candidates for a certain position. At present, there are different professional societies for biochemistry and molecular biology (PSBMB), cell biology (PSCB), developmental biology (PSDB), biology teachers (BIOTA Philippines), systematic biologists (ASBP), freshwater sciences (PSFS), and biodiversity and conservation (BCSP), to name a few. None of these organizations offer professional certificate programs for their members, save perhaps for the usual conference, training course, or seminar-workshop that they offer on an annual basis.

If we would be able to come up with a united front, we can be assured that we will be able to tap into this rich resource of potential contributors to help improve our awareness and understanding of the natural world, which, given this state of the new normal, should be integral components of our workforce the biologists. Rappler.com

Rey Donne S. Papa holds a PhD Biological Sciences degree from the University of Santo Tomas (UST), where he also holds the rank of Professor, teaching zoology and ecology courses in the UST Department of Biological Sciences. He is currently serving as the Dean of the UST College of Science and is the Program Lead for the Natural Sciences in the UST Graduate School. He is also the President of the Association of Systematic Biologists of the Philippines and the Vice President of the Philippine Society for Freshwater Science. Dr Papa is a freshwater biologist with 58 peer-reviewed international and national publications in the field to date. He may be reached through rspapa@ust.edu.ph.

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[OPINION] We need to professionalize biology in the Philippines - Rappler

Cell Biology

Hamilton Thorne Reports Record Revenue for the Quarter and Year-Ended December 31, 2019 – GlobeNewswire

BEVERLY, Mass. and TORONTO, April 22, 2020 (GLOBE NEWSWIRE) -- Hamilton Thorne Ltd. (TSX-V: HTL), a leading provider of precision instruments, consumables, software and services to the Assisted Reproductive Technologies (ART), research, and cell biology markets, today reported audited financial results for the fourth quarter and year-ended December 31, 2019.

Financial Highlights

David Wolf, President and Chief Executive Officer of Hamilton Thorne Ltd., commented, 2019 was another significant year for Hamilton Thorne with record sales of $35.4 million and adjusted EBITDA of $7.1 million. We continued to invest in sales and support resources in the US and Germany and continued to enhance our operations in order to take better advantage of the cross-selling and marketing synergies between the North American and European-based businesses. We also completed a significant expansion of our product line, geographic coverage and scale when we acquired UK-based Planer Limited in August. As we move past the COVID-19 pandemic, we see a significant opportunity to grow revenues from the Planer product line by leveraging our established direct sales channels and expanding the business of other Hamilton Thorne brands through additional direct sales capabilities in the UK.

Commenting on the quarter, Mr. Wolf added, With sales of $10.8 million and adjusted EBITDA of approximately $2.2 million, this was a record quarter for Hamilton Thorne. We increased our EBITDA margin versus the rest of the year, while continuing to invest in R&D and sales and marketing resources. Sales were positively impacted by the addition of approximately $1.6 million of revenues from the newly acquired Planer business, significant growth in the Companys laser and image analysis systems business, increased consumables sales, and continued growth of the sale of third-party equipment in the US. Gross profit for the quarter was up versus the balance of the year at approximately 56.5% due to product mix, and relatively flat compared to the prior years fourth quarter.

All amounts are in US dollars, unless specified otherwise, and results, with the exception of Adjusted EBITDA, are expressed in accordance with the International Financial Reporting Standards ("IFRS").

Results of Operations for the year-ended December 31, 2019

Hamilton Thorne sales increased 21% to $35,358,409 for the year-ended December 31, 2019, an increase of $6,144,595 from $29,213,814 during the previous year. Sales were up due to organic growth in operations owned by the Company for more than one year augmented by the added revenues from the Planer acquisition. Sales into the human clinical market grew substantially, primarily driven by strong increases in the sales of third-party equipment, our own clinical instruments, particularly in the second half of the year, our newly introduced LYKOS DTSTM laser system, and Gynemed-branded consumables, augmented by the contribution from the Planer acquisition. Sales into the animal breeding markets were up slightly for the year while sales into the research and cell biology markets were up substantially, largely driven by the contribution from the Planer acquisition as well as strong toxicology systems sales in the fourth quarter.

Gross profit for the year increased 15% or $2,526,302 to $19,030,022 in the year-ended December 31, 2019, compared to $16,503,720 in the previous year, primarily as a function of sales growth. Gross profit as a percentage of sales was down at 53.8% for the year-ended December 31, 2019 versus 56.5% for 2018 primarily due to product mix, particularly the impact of additional direct sales of third-party products in the US, and the addition of somewhat lower margin sales of Planer products, partially offset by increases in direct sales of higher margin proprietary equipment, branded consumables, and quality control testing services sales.

Operating expenses increased 22% or $2,897,173 to $15,805,307 for the year-ended December 31, 2019, up from $12,908,134 for the previous year, primarily due to the addition of Planer expenses post-closing, increased acquisition expenses, and increased depreciation and amortization. Excluding acquisition-related expenses for both periods, operating expenses would have been up $2,272,174, or 18%. Operating expenses were also affected by continued strategic investments in research and development and sales and marketing resources.

Net interest expense decreased $47,233 from $1,151,455 to $ 1,104,222 for the year-ended December 31, 2019 versus the prior year primarily due to reductions in the Companys revolving line of credit, convertible debentures due to conversion, and other term loan borrowings, plus interest earned on the Companys cash balances, partially offset by increased term debt in August to partially finance the Planer acquisition.

The change in fair value of derivative decreased $846,199 for the year-ended December 31, 2019, from a gain of $572,621 in 2018 to a non-cash loss of $273,578, primarily due to the weakening of the euro, partially offset by the increase of the Companys share price between the measurement dates.

Net income decreased to $793,275 for the year-ended December 31, 2019, versus $2,960,355 for the prior year, primarily attributable to increased revenues and profitability for the relevant periods offset by increased operating expenses, the $846,199 change in fair value of derivative, increased income taxes, increased acquisition expense, and continued strategic investments in research and development and sales and marketing resources.

Adjusted EBITDA for the year-ended December 31, 2019 increased 15% to $7,096,012 versus $6,187,254 in the prior year, due to revenue and gross profit growth, partially offset by increased operating expenses in the periods.

Results of Operations for the Fourth Quarter ended December 31, 2019

For the three months ended December 31, 2019, sales were up 34% from $8,072,739 to $10,841,097. Gross profit was up 34% to $6,121,965 versus $4,574,835 for the prior year. Sales and gross profit were up due to organic growth in operations owned by the Company for more than one year augmented by the added revenues and gross profit from the Planer acquisition. Gross profit percentage decreased from 56.7% to 56.5% for the quarter, primarily due to product mix. Operating expenses were up 34% to $5,160,706 versus $3,842,854 for the prior year, primarily due to the addition of Planer operating expenses for the quarter.

In the fourth quarter of 2019, the Companys net income decreased to $930,581 while Adjusted EBITDA increased 28% to $2,233,064 versus net income of $2,745,020 and Adjusted EBITDA of $1,750,245 for the prior year fourth quarter. These changes were due primarily, in the case of net income, to the $939,562 change in the fair value of derivative and increased income taxes, and in the case of Adjusted EBITDA to increased sales and gross profits partially offset by increased operating expenses.

See the Companys published Management Discussion and Analysis for a reconciliation of Adjusted EBITDA to Net Income for the year and the quarter.

OUTLOOK

The recent outbreak of the coronavirus, or COVID-19, has added substantial uncertainty to the short- and mid-term outlook as the countries where the Company has significant operations have required entities to limit or suspend business operations and have implemented travel restrictions and quarantine measures.The Companys operations are deemed to be part of the essential medical infrastructure in most places where it has personnel and it has implemented business continuity plans, including work from home programs, to maintain operations.

While it is not possible at this time to estimate the impact that COVID-19 could have on the Company, the continued spread of COVID-19 and the measures taken by the governments of countries affected has and is expected to continue to affect the demand for certain of the Companys products and services and could disrupt the supply chain and the manufacture or shipment of product inventories and adversely impact the Companys business, financial condition or results of operations. The COVID-19 outbreak and mitigation measures may also have an adverse impact on global economic conditions which could have an adverse effect on the Companys business and financial condition. The extent to which the COVID-19 outbreak impacts the Companys results will depend on future developments that are highly uncertain and cannot be predicted, including new information that may emerge concerning the severity of the virus and the actions to contain its impact.

Mr. Wolf commented, The first quarter of 2020 started off very strong. Our preliminary estimated results for Q1 are approximately $10.2 million in sales, which represents over 30% year over year growth and approximately 10% organic growth (approximately 12% constant currency). We did, however, see reduced demand for some of our products and services in certain territories as the quarter developed, as many IVF clinics reduced their activities. This reduction has continued in the second quarter, but we have also seen demand for our products return in some territories, notably China, which had been severely impacted in Q1. These fluctuations in demand for many of our products and services will last for a period of time that is difficult to determine, and will have an adverse effect on financial results in the second quarter, which could extend for one or more subsequent quarters. To improve our financial results, we have taken actions to reduce many non-essential expenses, reduce some personnel costs, and defer certain capital expenditures and new hiring.

Mr. Wolf continued, The Company has generated cash from operations since 2013 and, in normal circumstances, expected to generate cash from operations in 2020. Given the uncertainties surrounding the COVID-19 outbreak, it is impossible to predict whether the Company will generate cash from operations in 2020. Regardless, the Company currently maintains a strong balance sheet, with cash on hand of approximately $15.1 million, and net bank debt of approximately $10.1 million following the Companys Q1 drawdown under its line of credit, and believes that its current cash position should be sufficient to support operations for the next twelve months. We continue to work on our acquisition program with a goal of completing one or more meaningful acquisitions every twelve to eighteen months; however, the effects of the COVID-19 outbreak could affect this goal.

Mr. Wolf added, The COVID-19 virus will eventually run its course, and, we remain optimistic that once this happens, the strong macroeconomic and demographic tailwinds that have driven the growth of our business over the past few years will continue for the foreseeable future.

Conference Call

The Company will hold a conference call on Wednesday, April 22, 2020 at 11:00 a.m. EDT to review highlights of results. All interested parties are welcome to join the conference call by dialing toll free 1-855-223-7309 in North America, or 647-788-4929 from other locations, and requesting Conference ID 7281879. The Companys updated investor presentation and a recording of the call will be available on Hamilton Thornes website shortly after the call.

Financial Statements and accompanying Management Discussion and Analysis for the periods are available on http://www.sedar.com and the Hamilton Thorne website.

About Hamilton Thorne Ltd. (www.hamiltonthorne.ltd)

Hamilton Thorne is a leading global provider of precision instruments, consumables, software and services that reduce cost, increase productivity, improve results and enable breakthroughs in Assisted Reproductive Technologies (ART), research, and cell biology markets. Hamilton Thorne markets its products and services under the Hamilton Thorne, Gynemed, Planer, and Embryotech Laboratories brands, through its growing sales force and distributors worldwide. Hamilton Thornes customer base consists of fertility clinics, university research centers, animal breeding facilities, pharmaceutical companies, biotechnology companies, and other commercial and academic research establishments.

Neither the TSX Venture Exchange, nor its regulation services provider (as that term is defined in the policies of the exchange), accepts responsibility for the adequacy or accuracy of this release.

The Company has included earnings before interest, income taxes, depreciation, amortization, share-based compensation expense, changes in fair value of derivatives and identified acquisition costs related to completed transactions (Adjusted EBITDA) as a non-IFRS measure, which is used by management as a measure of financial performance. See section entitled Use of Non-IFRS Measures and Results of Operations in the Companys Management Discussion and Analysis for the periods covered for further information and a reconciliation of Adjusted EBITDA to Net Income.

Certain information in this press release may contain forward-looking statements. This information is based on current expectations that are subject to significant risks and uncertainties that are difficult to predict. Actual results might differ materially from results suggested in any forward-looking statements. The Company assumes no obligation to update the forward-looking statements, or to update the reasons why actual results could differ from those reflected in the forward-looking statements unless and until required by securities laws applicable to the Company. Additional information identifying risks and uncertainties is contained in filings by the Company with the Canadian securities regulators, which filings are available at http://www.sedar.com.

For more information, please contact:

David Wolf, President & CEO Hamilton Thorne Ltd. 978-921-2050 ir@hamiltonthorne.ltd

Glen AkselrodBristol Investor Relations905-326-1888glen@bristolir.com

Michael Bruns, CFOHamilton Thorne Ltd.978-921-2050ir@hamiltonthorne.ltd

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Hamilton Thorne Reports Record Revenue for the Quarter and Year-Ended December 31, 2019 - GlobeNewswire

Cell Biology

Male reproductive health problems could begin in womb – Mirage News

Disruptions to male babies development early during pregnancy could have a profound effect on a mans future reproductive health, according to new research.

A world-first finding into the gestation period when testes develop in the fetus has given researchers new insights into the potential causes of male infertility and testicular cancer later in life.

A Hudson Institute research team led by Professor Kate Loveland has discovered that a growth factor, activin A, is linked to steroid production in the testes in utero.

This research demonstrates for the first time that activin A is necessary for normal production of testosterone in the fetal testis, said Ms Whiley, a PhD student with the Testis Development and Male Germ Cell Biology group and the studys first author.

Published in the journal Endocrinology, the discovery shows that activin A promotes synthesis of two enzymes crucial for the final steps of testosterone synthesis, in a preclinical model. However, the absence of activin A resulted in an abnormal steroid environment.

Ms Whiley said the findings pinpoint an important growth period in utero when the testes are developing.

Events that alter activin A levels, which can occur due to different physiological conditions of pregnancy or the mothers exposure to certain medications, may explain why some boys and men have impaired reproductive health, Prof Loveland said.

Testicular cancer is the second most common cancer in young men aged 18-39. Reduced fertility affects one in 20 men. Both are increasingly common, but the causes of these disorders are not well understood. Disruptions occurring during fetal life may have a profound effect on a mans future reproductive health, she said.

The research provides a new understanding of how steroids are produced in the fetal testis. In addition, the key role of activin A highlights how formation of the earliest germline cells may be affected by factors in their environment.

This study is part of our ongoing research that aims to better understand the basis of male infertility and conditions such as hypospadias, cryptorchidism (see Key facts below) and testicular cancer, which are increasing worldwide, Prof Loveland said.

KEY FACTS

Funders | NHMRC

Collaborators | Professor David Handelsman (ANZAC Institute, Sydney, AUS), Professor Kristian Almstrup (Copenhagen University Hospital)

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Male reproductive health problems could begin in womb - Mirage News

Cell Biology

Accelerating Antibody Discovery with Next-Generation Single-Cell Analysis – News-Medical.net

Why are therapeutic antibodies traditionally produced in vivo during early drug discovery?

Typically, in vivo models are used because they are cost-effective and relatively fast at generating candidate antibodies that can be tested. Introduced by Khler and Milstein in 1975, the traditional mouse hybridoma technique begins by injecting a validated or known antigen into the animal model, for example, mice, which causes an immune response that generates novel antibodies. These novel antibodies can then be harvested and fully characterized downstream to ensure that they have the desired molecular function and properties.

Image credit: extender_01 | Shutterstock

Scientific and technological advances have further improved in vivo studies. More recently, humanized mice have been immunized to express human immunoglobulins, rather than the mouse antibody genes, which enhances how translational these are downstream.

Animal models continue to be prevalent, primarily because they are tried and tested in the industry with good results. Despite the challenges, hybridoma technology is a well-established platform, and the humanization of animal models has further promoted this method. Most FDA approved therapeutics have been produced using humanized platforms.

However, fully rodent animal models can result in rodent specific immunogenicity and proteins from the animal model may remain at a later stage. In fact, not all antibodies produced in humanized animal models will be entirely human.

Other technologies, for instance, phage display provide a compelling alternative as they dont require animal models, have shorter timelines and researchers can use a fully human antibody library. These libraries can be quite diverse and can contain different constructs within them, for example, single-domain antibodies. Using next generation sequencing (NGS) with phage display helps characterize immunoglobulin genes from large populations, which better represents natural human antibodies.

Phage display isnt without downsides either; as it is a prokaryotic system rather than mammalian, there may be issues with affinity and other properties of this in vitro platform.

After successfully immunizing an animal, a lymphoid organ, for example, the spleen, is harvested. This is processed and B-cells are isolated. B-cells that produce antigen-specific antibodies that bind to the target antigen then need to be separated from those that bind to other antigens.

Next, the reactivity to the target antigen is tested to ensure that selected antibodies bind specifically to the antigen (rather than to a range of antigens) and that this affinity and sensitivity is above the required level. Once the unique B-cells have been isolated, scientists will screen for binding characteristics, cross-reactivity and more.

Research teams will typically use flow cytometry to screen the B-cells directly or more often produce hybridomas which are then screened.

Producing a hybridoma population requires a fusion reaction between the B-cell and a cancerous myeloma cell. This allows the B-cell to proliferate and survive indefinitely in culture, which is essential to safeguard potential cells of interest. Unfortunately, when creating hybridomas, the whole B-cell population is unable to be fused; therefore, only a fraction of the total population can be screened, thus leading to the loss of a large part of the B-cell repertoire.

Traditionally, flow cytometry is favored because of its high-throughput, and antibodies secreted from the B-cells can potentially be screened using cold capture, a technique used to manipulate the cell to prevent the full secretion of antibodies by trapping them at the cell surface. However, this is a representation rather than direct measurement of antibody secretion.

Alternative screening methods include ELISA and Elispot, which can measure B-cell secretion. Although these techniques often need to be completed manually, making them tedious and limited to low-scale screening.

A research team typically spends many months during this process cycle, from immunizing the animal model, collecting cells and creating hybridomas or directly screening B-cells. The population of target cells can be as low as 0.001% of the original ~40 million cells in the harvested organ. Further loss due to diminishing cell viability and cell death due to the harsh processing of these cells can severely impact productivity.

After completing this cycle, the target cells may not be found because they have either not been successfully fused during the hybridoma fusion process or died during culture due to the harsh processing. This is a big challenge in the industry.

The Cyto-Mine Single Cell Analysis System can overcome numerous challenges in the industry. First, the need for high-throughput yet gentle technology that enables the deep interrogation of whole B-cell repertoires and hybridoma populations while preserving cell viability. Second, the need for highly sensitive and specific assays to find rare antibody-secreting cells with the desired antibody characteristics.

Cyto-Mine can be used in screening entire cell populations to find that rare B-cell or hybridoma that secretes the antigen-specific antibody. In fact, researchers can input isolated B-cells directly into the Cyto-Mine, which drastically cuts down the process cycle timelines, and removes the possibility of losing cells in culture. Additionally, Cyto-Mine offers the flexibility in assay design, of which there is a great need due to the varying nature of the targets and the biologics.

Underpinned by Sphere Fluidics patented picodroplet technology, Cyto-Mine integrates the selective screening of tens of millions of single cells in miniaturized aqueous picoliter compartments called picodroplets.

Picodroplets provide individual micro-reaction chambers where secreted proteins accumulate and can be assayed to identify valuable proteins (e.g. antibodies with antigen-specificity), to find rare cells of interest or to find high-producing cells.

The picodroplets themselves provide a protective environment for the fragile B-cells, shielding them from shear stress as the picodroplets travel through the microfluidic channels. Cells can also be encapsulated in their preferred culture media, so there are no harmful culture transitions.

By combining B-cell isolation and identification of antigen-specific hits into a process that takes only 1-2 days in Cyto-Mine, you significantly reduce the stress on the fragile cells and accelerate the steps to downstream analysis such as functional testing or sequencing.

Image credit: Sphere Fluidics

Users must know whether the antibody is an IgG or an IgM, before making detection probes that will bind to the target of interest. The assay format is based on a simple FRET reaction, and two detection probes must be encapsulated into the picodroplet along with the cells. One that binds to the Fc region of the antibody and another that targets the antigen on that antibody. Both probes are conjugated to a fluorophore, making a pair of detection probes that can induce a FRET reaction.

When the detection probes are in close proximity, i.e. if they have bound to the Fc and Fab regions of the secreted antibody, they will induce a FRET-mediated shift on fluorescence where the excitation of one probe will transfer energy to the other which will emit a fluorescent signal. Cyto-Mine detects for an energy transfer within the picodroplet, and as the detection probes are customized to target only the antigen of interest, there will not be a fluorescent energy transfer if the antigen of interest isnt present.

Cyto-Mine supports flexible assay design, so the assay format can be tailored to make it specific to the target.

Developed with biopharma in mind, the initial design of Cyto-Mine relied on input from key pharma companies, to solve their most significant challenges and automation was vital to do this.

The current process is incredibly time-consuming and tedious. New techniques need to have a high-throughput, diversity and flexibility of the assay. Rather than automating pipetting, weve made an entirely novel system to approach antibody discovery.

The Cyto-Mine is also applicable to cell line development, where there is a similar need to automate inefficient processes, and Cyto-Mine has single cell imaging and dispensing capacity making it a great fit.

Theres an incredibly high demand for antibody-based biotherapeutics, and its easy to see how successful they are with over 500 antibody-based therapeutics beginning to come through clinical trials. This demand is exemplified with one of the Antibody Societys 20 antibodies to watch in 2020, an antibody against the Ebola virus. The company producing this antibody could stop trials before completion because it was so successful compared to any other treatments.

Over the next decade, the antibody discovery process needs to become quicker, and as new technology is incorporated, should become more rapid. This optimization will shorten the time to market and get new therapeutics in the hands of patients faster.

Dr Zoe Nilsson is the Global Product Marketing Manager for Sphere Fluidics. Zoe has a BSc, MPhil and PhD in Neuroscience and Cell Biology from the University of St Andrews, UK. Zoe works closely with the R&D team to develop new features of Cyto-Mine and novel assay reagent kits to help biopharma customers identify and isolate rare cells of interest.

Zoe is an experienced Product Manager with a demonstrated history of working in the biotechnology industry. Before joining Sphere Fluidics, Zoe led the development of novel cell types from induced pluripotent stem cells (iPSCs) for use in basic research, drug discovery and safety toxicity testing.

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Accelerating Antibody Discovery with Next-Generation Single-Cell Analysis - News-Medical.net