Re: [DIYbio] Re: How safe is lysed E.coli as an adjuvant?

Just to change direction here a bit, (because in principle I do want to see better knowledge-bases on how to do medicine far from privilege): what's wrong with using inorganic adjuvents, which probably have a safer dose/response curve, probably lower response variability, and might be easier to determine purity/sterility?

On 10 March 2020 09:24:38 GMT+00:00, Michael Flynn <mfly...@gmail.com> wrote:

For the record, I agree with you both that a random person shouldn't just try to "DIY it".

However, I think some of the risks you mentioned can be attacked. Humans are very sensitive to LPS, with a fatal dose ranging in 1 ug/kg, however people have survived (and ethics boards have cleared experiments using!) doses of 4 ng of *purified* LPS per kg, so around 140 ng of LPS (https://www.ncbi.nlm.nih.gov/pubmed/2835680/). Assuming a saturated culture is completely made up of E.coli, about .1 ul of that culture would come out to around 100 ng of E.coli and thus would be around that dose, assuming E.coli is entirely made up of LPS, which it is not. At the same time, .1 ul of saturated culture would probably contain enough E.coli to mount an adaptive response against. You could even dilute further down to .01 ul... if an e.coli is about 1 femtoliter then .01 ul still contains 10 million e.coli so...  seems like it would be enough. Also I'd like to reiterate that these would be dead E.coli that have been thermally lysed. 

Anyways, at that point you wouldn't be dying of LPS toxicity. And the idea would be to proceed by eliminating risks one by one until the risk/reward payoff is positive, even if the payoff is small (slight chance of becoming immune to virus that probably won't kill you). 

On Monday, March 9, 2020 at 2:30:44 PM UTC-7, Andreas "Mega" Stuermer wrote:

I think I read in a paper, long ago, that LPS contamination might contribute to DNA vaccine immunogenicity. 

I would assume that expressing the protein in coli is not ideal - you get so little protein vs so much LPS. It might just cause the innate immune system to go bonker. Our immune system is trained to respond to the tiniest ammounts of LPS. Also you will get a B-cell response this way, but not much T-cell response. De-novo synthesis of proteins and display of fragments thereof mediates ADCC activity and the lack thereof is (for example) the reason why Herpes vaccines have failed so far. 

I'm not often one to say this isn't DIY-ble but considering everything the danger seems very high compared to potential gains. Sure, given enough training you could turn your basement into a certified 50k$ lab that is suitable. May be easier to join a lab. 

As far as I can tell, live recombinant or live attenuated viruses are the new thing for effective vaccines - here they inserted an Ebola protein into a "harmless-to-human" virus that replicates https://www.nejm.org/doi/full/10.1056/NEJMoa1414216 

But then, there's a lot of regulation around that topic and it is not smart to play around with this in a DIY setting. 

Sometimes there's a reason why things are mostly done in academia.

On Saturday, March 7, 2020 at 3:21:32 AM UTC+1, Michael Flynn wrote:

Often in vaccine development, in addition to target antigens that you want to raise antibodies against, an adjuvant, something that stimulates the immune system like lipopolysaccharide, must be added in order to generate an adaptive immune response against the antigen. E.coli is covered with lipopolysaccharide, which reacts strongly to complement (antibacterial factors in blood), and so would be a good candidate for an adjuvant. This line of thinking leads to an "easy vaccine" where you express the antigen in E.coli and thermally lyse the cells, dilute them to some concentration in water for injection, and there you have your vaccine. In an outbreak like coronavirus, making vaccines like this would seem to be "low risk, high reward" in that nothing bad seems likely to come of it, but you could potentially immunize against this bad outbreak.

Anyone have any actual literature on this?

--
Sent from my Android device with K-9 Mail. Please excuse my brevity.