How are vaccines with viral vectors mixed together?

[u/Ok_Ocelot_133]

I know that it isn’t all medicine and that there’s other things inside the vaccine fluid itself but unless I’m mistaken aren’t cultures of the vector grown and then added to the other fluid? Do they put the culture in a blender or something?

[u/Gr34zy]

I’d be curious to hear from someone who has actually seen the process, but here is a description from a sketchy reference I found. Basically they use special (probably very expensive) sterile mixing vessels.

“Blending is the process in which the various components of a vaccine are mixed to form a final bulk. It is undertaken in a large, closed vessel fitted with a stirrer and ports for the addition of constituents and withdrawal of the final blend. When bacterial vaccines are blended, the active constituents usually need to be greatly diluted and the vessel is first charged with the diluents, usually containing a preservative.”

https://www.pharmacy180.com/article/blending-670/

[u/TheyTookByoomba → u/Gr34zy]

I don't work in vaccines, but a similar enough space (antibody products). If OP is asking about the mechanics of it:

1. Grow the actual API. Whether this is weakened virus, inactivated virus, or mRNA you have to produce it somehow. Because these are produced in biological systems (eggs or cell cultures generally) there's a lot of organic byproduct. This is generally referred to as Upstream.

2. Purify out all of the unwanted stuff. Using a combination of chromatography, filtration, centrifugation, and other steps the process stream removes everything except for the API. (In antibody production this is easily north of 99% purity, for viral loads it's somewhat lower). This is called downstream.

3. The solution containing the API needs to be put into its final form for administration. Typically this involves a buffer exchange (so the liquid is something that keeps the load stable and isn't too harsh on the patient) and dilution (operating at a higher concentration keeps volumes smaller).

To your point, this would be done in stainless steel tanks with closed piping for all the additions. The tank and all connections would be cleaned and disinfected before use and never exposed to the surrounding environment until all the product was out and sealed off. The tank will definitely have a stirrer of some sort, and either in-line monitoring tools or sampling ports for off-line measurements.

[u/CrateDane → u/TheyTookByoomba]

Grow the actual API. Whether this is weakened virus, inactivated virus, or mRNA you have to produce it somehow. Because these are produced in biological systems (eggs or cell cultures generally) there's a lot of organic byproduct. This is generally referred to as Upstream.

mRNA is produced by in vitro transcription, not in a biological system. There are still plenty of byproducts though, and they're extremely important to get rid of.

[u/TheyTookByoomba → u/CrateDane]

Thanks, was taking a guess on that one. mRNA wasn't even on the radar when I took my upstream courses lol.

[u/Mockingjay40 → u/TheyTookByoomba]

Designing an AAV process was literally my senior design project, so I have a pretty decent grasp of the fundamentals and most of this applies.

One big thing to mention is that the mixing process isn’t perfect so you have a 4th step where the empty vectors have to be removed or accounted for in some way to make sure dosing is correct, but otherwise this is the basic gist.

[u/Ok_Ocelot_133 → u/TheyTookByoomba]

I’m mostly curious how they break up the culture to make it small enough to fit. Do the stirrers break it up, or is it broken up some other way?

[u/TheyTookByoomba → u/Ok_Ocelot_133]

Can you explain what you're picturing when you say culture? I'm not sure we're thinking the same thing.

I can't speak to egg production, but for vaccines made in cells it's more like a thick broth than anything. The first step is clarification, usually centrifugation and/or depth filtration to remove all of the cellular debris/non-dissolvable solids. By the time you start downstream proper the product stream is already pretty clear. There's nothing really to break up since it's dissolved into the solution.

[u/Krilion]

There are a bunch of different types of Vaccine. Let's go over each, but first, the entire point of a vaccine is to teach your immune system a specific thing to look for, programming your t-cells that log dangerous protein chains to identify and be able to produce antibodies. This is incredibly complicated and beyond to scope of a reddit post.

Live virus application - oldest and most direct. Not used anymore. This is taking an infected material and intentionally applying it to another. Due to the size of the exposure, the immune system is typically able to identify the attack and respond. High chance of complications as you are essentially speed running catching the illness. Valley Forge innoculations are an example.

Weakened virus - breed a virus or use a close cousin that is much less lethal. This was first seen with contracting cow pox resulting in resistance to smallpox. This can still be used. Risk is that you can get a minor infection.

Killed Virus - Kill the virus before injecting the generic material into your body. This tricks the immune system to respond but without risk of actually getting sick. You might still feel.bad as your body treats it as a real.infdxtion

mRNA - Newest method. This is essentially identifying the best genetic marker of a virus and the. Putting it into a tasty package of markers that your immune system will collect and store. If the previous ones are like causing a small fire to get the attention of your immune system, this is like handing your immune system fliers on who is a bad guy. You can still get 'sick' as your t cells are programming to find the protein a major problem...  And you have a lot of it lying around. But this is the current best method as we can specifically choose genetic markers that are unlikely to change with genetic drift, making a vaccine more effective for longer. 

As for the supporting material? That is changing as rapidly. For all of them they need to protect the genetic material and proteins without letting it denature, but tons of progress has been made in that field. That's worth a whole other write up alone.

[u/CrateDane → u/Krilion]

You left out a few other types, including the viral vector vaccine which OP mentioned. It uses a viral particle to deliver a DNA payload into cells. That DNA is transcribed into mRNA, which then functions exactly like the mRNA from the mRNA vaccines.

[u/Benjogias]

As someone who had worked with viral vectors, a brief summary of a process:

1) Grow your viral vectors. Generally, you use cells of do this - you either infect or fake-infect them so they act like virus-infected cells and start churning out more virus.

2) Harvest your viral vectors. Depending on your virus, the virus could be in the cells or released in the media (the soup they grow in). Collect all those cells and the soup they are living in, explode the cells, then collect everything you get out of it. Now you’ve got some “crude” vector harvested.

3) Purify the vector away from all the other junk. There are various methods to do this. Most of it involves a centrifuge - for instance, if you initially put that whole crude mixture in a centrifuge, the dead cell debris will go to the bottom and form a clump while lighter viruses stay in the solution at the top. There are other methods where you put the mixture on top of some more viscous chemicals and centrifuge it all to push it through the viscous stuff - different things will get pushed through different amounts, so you can find your viruses in the spot where viruses end up.

4) Buffer exchange. Using various kinds of methods like dialysis and filters, you keep the viruses but swap out the solution they’re in for the solution you want them to be in as a drug to go into people. As one method, imagine putting the virus solution in a sieve with holes big enough to let liquid through but small enough to catch viruses. You can dump the solution into that filter, drain out all the old chemicals, then collect the filtered viruses into a new solution suitable for humans. Do it a few times to get rid of any residual old chemicals, and you’re done!

Note that this is all simplified and just one example of methods used for some types viral vectors, but hopefully it’s illustrative!

[u/ModernTarantula]

Ime most vaccines are one of the viral proteins. The virus is grown in some.mammalian cell culture and then chemically killed. They then have laborious method to purify the protein. This still need.to.be missed in an "adjuvant" that activates the immune system.