More lot surveillance reveals DNA contamination variance

Over the Holidays, we received some more vaccines from various collaborators.

One lot that stood out was BioNtech 1F1042A. This had the lowest CT score ever recorded on a lot and currently holds the most contaminated record.

You can see IF1042A clocking in at a CT of 12.72 with SV40. This is 4-5 CTs (2^5 = 32 fold) more DNA than other vaccines tested to date. These are vaccines that Ulrike Kaemmerer is applying to cancer lines. The readings above are before the application to the cell lines listed in their names so they are direct readings from the vaccine vials. She then passages them and we track the vaccine CTs scores with each passage and whole genome sequence interesting events.

This is a reminder that the majority of the DNA contamination studies performed to date are across a narrow number of actual lot numbers. As this is expanded to more lot numbers we should not be surprised to find 16-32 fold changes in DNA contamination as the EMA witnessed 211 fold variances.

Since we have these reagents out, I decided to burn the remainder of the kit on other vials that were sent to us anonymously over time. These make for good controls when your’e working with vaccinated cell lines. Its important to have pure vaccine to run side by side with any cell lines as a positive control for your qPCR/RT-qPCR. These originate from Japan. 7 Vials in total from 5 lots.

Lot Numbers and Vials

Lots from Japan

SV40 isn’t in every vaccine so its not surprising to get no SV40 signal for the Spikevax samples. The more recent XBB vaccines also contain DNA contamination. These don’t expire until 2025. As you can see expired or non-expired vials both have DNA contamination and both were injected into people. The Strawman arguments they erect should really be called Stringmans as they are so thin and easily discarded.

Samples were run in triplicate at 1:10 dilution and Neat (No dilution). The Neat samples often show qPCR inhibition thus 1:10 dilutions are recommended and adjustment (Subtraction) of these CT score with 3.3CT to account for the initial CT score.

The First dilution in the standard curve is 400pg (4ng into 50ul ddH20 and 5ul (1/10th) of this being used in qPCR). The 3rd dilution is where these vaccines are clocking in on the 4pg standard. The plasmid is approximately 74X larger than the amplicon (296pg/ul). 300ul injections = 88.8ng/dose.

To cross check this with samples run in previous studies…

Speicher et al highest sample was 4.3ng/dose with qPCR CT of 18.44 on Ori and a SV40 CT of 16.99. This is a delta CT (between Speicher et al SV40 signal on the highest lot and the SV40 on this lot) of 16.96-12.72 = 4.18. 2^4.18 should equate to 18.13X more DNA than the highest sample published in Speicher et al. This would be 77.3ng/ul according to his standard curve.

A fluorometry reading has not been performed yet on this vaccine but those typically are 10X higher in quantitation as they can detect DNA smaller than 105bp.