The Human Adenovirus Type 5 Reference Material (ATCC VR-1516) is now available. All new stocks of the reference material will be produced and authenticated at ATCC instead of by an outside working group—that means no more supply limitations when ordering!
Because of the necessary differences in manufacturing between new lots and the original 2001 production, there are a few changes to the reference material as compared to previous stocks. We have provided a comparison below for reference.
Table 1. Culture comparison of Human Adenovirus Type 5 reference material produced by ARMWG and ATCC
| ARMWG-produced lots | ATCC-produced lots | |
|---|---|---|
| Description | Purified Human Adenovirus Type 5 Reference Material | Purified Human Adenovirus Type 5 Reference Material |
| Backed by working group | Yes | Yes |
| Origin | Master virus bank derived from plaque-purified Human adenovirus 5 (ATCC VR-5; wild type) | Master virus bank derived from plaque-purified Human adenovirus 5 (ATCC VR-5; wild type) |
| Host cells | HEK 293 cells | HEK 293 cells |
| Particle concentration | 5.8×1011 particles/mL, with 95% certainty that the range lies within 5.6×1011 to 6.0×1011 particles/mL, as determined by A260 nm/SDS procedure, based on data submitted from 13 different laboratories. | 1.03×1011 particles/mL using a modified method that provided more consistent results |
| Genome copy number | Not performed | 1.41×109 genome equivalents per mL as determined by ddPCR™ in triplicate with 9 replicates |
| Infectious titer (NIU/mL) | 7×1010 NIU/mL in HEK 293 cells, with 95% certainty that the range lies within 7×1010 to 8×1010 NIU/mL, based on data submitted from 17 different laboratories. | 1.27×1012 NIU/mL in HEK 293 cells |
| Infectious titer (TCID50/mL) | TCID50/mL not reported | 5.00×1011 TCID50/mL |
| Electron microscopy | Confirmed >70% of the material consists of icosahedral shaped single adenovirus particles, with the remaining population being doublets, triplets or multiplets consisting of more than three adenovirus particles. | Not performed |
| Authentication | The complete DNA sequence was determined and is deposited with GenBank. The ARM GenBank accession number is AY339865. The sequence is similar, but not identical, to the adenovirus 5 sequence found in GenBank accession number NC_001406 (Human adenovirus 5, complete genome). | Next-generation sequencing was performed. The new lot was compared with the reference sequence and the original lot to verify the sequence and identify nucleotide variants to the reference sequence (AY339865.1). |
| Adventitious agents | Tested and found free of adventitious agents |
Tested and found free of adventitious agents |
| Mycoplasma (direct) |
Not reported separately | None detected |
| Mycoplasma (PCR-based) | Not reported separately | None detected |
| Sterility | Not reported separately | No growth |
| Endotoxin | Endotoxin was < 0.15 EU/mL | Endotoxin was < 0.15 EU/mL |
| Residuals | Less than 3 pg of residual host cell DNA per µg of total DNA. Residual HEK 293 host cell proteins were found to be 18 ng/mL. Residual BSA was found to be <0.5 ng/mL. There is approximately 2.0 µg of free hexon per 1012 adenoviral particles. No 31K MW precursor protein form was found. The A260 nm to A280nm ratio in the presence of 0.1% (w/v/) SDS was 1.37. |
Residuals – RnaseP ddPCR – Residual host cell DNA below the limit of detection The A260 nm to A280 nm ratio in the presence of 0.1% (w/v) SDS was 1.81 |
| Bioinformatics analysis |
Not performed | Adenovirus variants will be detected relative to the following genome as a reference sequence: AY339865.1 (Adenovirus) from GenBank. |
| Fill volume | 0.5 mL | 0.5 mL |
| Storage | -70°C or colder | -70°C or colder |
| pH | 8.0 at room temperature | 7.5 post-thaw (using a pH strip) |
| Manufacturing standard | GMP | ISO 9001 |
NIU = NAS infectious units
NAS = normalized adjusted standard
NGS = next generation sequencing
BSA = Bovine serum albumin
ARMWG = Adenovirus Reference Material Working Group
EU = Endotoxin units
