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I had the same question in the last video, but in that case he seemed to indicate he was testing for the terpene and related metabolic or degradation products of it. But yeah, knowing the potential for non-specific activation by any/all the possible real-world environmental elements, which could vary between samples... would be pretty high on my list of things expected necessary to convince VCs/investors/academics.
We get these kinds of questions from reviewers too. It will be addressed a bit in my next video going over PYR1 sensors in more detail. Unfortunately, it's impossible to test for cross reactivity of all chemicals! But we test as many relevant things as we can to prove our point that our sensors are specific
@@DIYBiotech thanks for your reply. Is it like how in covid test, based on the viral load the signal which is given is more intense in the antibody test? Also, just binge watched 10 of your videos now, really like your content.
Are there limits of detection for PYR1 biosensors? Is the downstream signal cascade dependent on ligand binding affinity? I guess I'm wondering if these biosensors are only capable of spitting out a binary yes or no response to the presence of some ligand. Or is there a way to somehow account for the degree to which GFP is expressed and then do a back of the envelope calculation for analyte concentration?
Great questions! So far we have some sensors that detect down to single nanomolar levels. The downstream cascade is independent of binding affinity. There's a bit on how these sensors can respond in my dissertation video and also in some videos that will come out in the next couple months! Short answer is that the linear range of these sensors can be one or two orders of magnitude.
Org. Chemist here, and I wonder what's the false negative rate on these sensors. Just from a combinatorics pov if we assume that it's mostly the binding residues that determine the binding to the ligand, that leaves us with somewhere around 20^10 options. That's still approx. 10^13 which is just over the number of small molecules accounted for in the chemical space project. So even undercounting the variability space of the sensors, it is bigger than the variability space of the signal that they are sensing (ligands) meaning one would expect just by chance that the same ligand would be sensed by multiple sensors, leading the false negatives. This is ofc a statistics argument for why false negs would be expected but how is this solved from a bioengineer's pov?
Just to clarify 20^10 cus 20 amino acid residues combined into 10 binding residues. Ofc not all combinations are valid, cus 20 glycines would ofc just make empty space, but just for an approximation. And 10 cus I've never seen a docking experiment with over 10 binding residues in the xrd structure
Fantastic thought. The main library we use only has certain mutations for each residue, resulting in a library of only 30,000 mutants. We then plate ~3 million cells, so that we have 100X coverage of the library. This way, we can statistically say there are very likely no false negatives. However, often we don't care if we miss a sensor or two, we just want to see if we can find any sensors. For this, having a library of millions or billions of mutants could still be useful. A more detailed PYR1 video is coming soon!
Hey man can you review this really interesting paper on frontiers. It's called Construction of an efficient Claviceps paspali cell factory for lysergic acid production. Would be amazing if you would make video of that. Really interesting study you did. And congratulations with your getting your diploma. I like your video styles it's really good education and you explain complex topics in an easy way.
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How do you control for false positives for environmental testing of pesticides using PYR1? are they that ligand-specific?
I had the same question in the last video, but in that case he seemed to indicate he was testing for the terpene and related metabolic or degradation products of it. But yeah, knowing the potential for non-specific activation by any/all the possible real-world environmental elements, which could vary between samples... would be pretty high on my list of things expected necessary to convince VCs/investors/academics.
We get these kinds of questions from reviewers too. It will be addressed a bit in my next video going over PYR1 sensors in more detail. Unfortunately, it's impossible to test for cross reactivity of all chemicals! But we test as many relevant things as we can to prove our point that our sensors are specific
@@DIYBiotech I'm sure NIST has an expensive library! haha...
This is super exciting research!!! I am a junior majoring in bioengineering and I hope one day this technology wide-spread, amazing!
Very interesting, testing for fent or zenes on the fly would be great
Great video my guy. You got yourself a new subscriber. I have a question, This method is a simple yes or no right. It's not a quantitative test ?
Thanks! They can be set up as binary, but many of our sensors have one or two orders of magnitude of linear sensitivity range.
@@DIYBiotech thanks for your reply. Is it like how in covid test, based on the viral load the signal which is given is more intense in the antibody test? Also, just binge watched 10 of your videos now, really like your content.
@@Itsnotfogell yea exactly like that! Thanks for the support bud!!
Sorry to pester. 😅 Can't we use the same method but in a microfluidic system so that it can act as a quantitative test?
@@Itsnotfogell I'm not too familiar with microfluidics, but from what I understand it could be used to increase the linear range of the sensor!
Are there limits of detection for PYR1 biosensors? Is the downstream signal cascade dependent on ligand binding affinity? I guess I'm wondering if these biosensors are only capable of spitting out a binary yes or no response to the presence of some ligand. Or is there a way to somehow account for the degree to which GFP is expressed and then do a back of the envelope calculation for analyte concentration?
Great questions!
So far we have some sensors that detect down to single nanomolar levels.
The downstream cascade is independent of binding affinity.
There's a bit on how these sensors can respond in my dissertation video and also in some videos that will come out in the next couple months! Short answer is that the linear range of these sensors can be one or two orders of magnitude.
Can u make the magic yeast at home?
Org. Chemist here, and I wonder what's the false negative rate on these sensors. Just from a combinatorics pov if we assume that it's mostly the binding residues that determine the binding to the ligand, that leaves us with somewhere around 20^10 options. That's still approx. 10^13 which is just over the number of small molecules accounted for in the chemical space project. So even undercounting the variability space of the sensors, it is bigger than the variability space of the signal that they are sensing (ligands) meaning one would expect just by chance that the same ligand would be sensed by multiple sensors, leading the false negatives. This is ofc a statistics argument for why false negs would be expected but how is this solved from a bioengineer's pov?
Just to clarify 20^10 cus 20 amino acid residues combined into 10 binding residues. Ofc not all combinations are valid, cus 20 glycines would ofc just make empty space, but just for an approximation. And 10 cus I've never seen a docking experiment with over 10 binding residues in the xrd structure
Fantastic thought. The main library we use only has certain mutations for each residue, resulting in a library of only 30,000 mutants. We then plate ~3 million cells, so that we have 100X coverage of the library. This way, we can statistically say there are very likely no false negatives.
However, often we don't care if we miss a sensor or two, we just want to see if we can find any sensors. For this, having a library of millions or billions of mutants could still be useful.
A more detailed PYR1 video is coming soon!
coool. thank you
Could make videos on the manufacturing of Ivermectin and Calcifediol?
Talking about biotech specifically id like to know more about BDNF and peptides personally
Whoa just looked it up, sounds cool!
Hey man can you review this really interesting paper on frontiers. It's called
Construction of an efficient Claviceps paspali cell factory for lysergic acid production.
Would be amazing if you would make video of that. Really interesting study you did. And congratulations with your getting your diploma.
I like your video styles it's really good education and you explain complex topics in an easy way.
Thanks for the encouragement! That paper looks really good so I'll have to do a video on it. DMT in bacteria is next month tho :) (to hold you over)
You should do a book review of blood music by greg bear
Thank you for your and your peers work. The world needs more greatness like this. Also lemme know when you get some of that special yeast🫡🤫