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Detection of contaminants in cell cultures, sera and trypsin
Tatiana Flávia Pinheiro de Oliveira1, Antônio Augusto Fonseca, Marcelo Fernandes Camargos
1Laboratório de Biologia Molecular/Laboratório de Diagnóstico de Doenças Virais, Laboratório Nacional Agropecuário de Minas Gerais, Pedro Leopoldo, Minas Gerais, Brazil.
Abstract:
The aim of this study was standardization and application of polymerase chain reaction (PCR) for the detection of contaminants in cell cultures, sera and trypsin. Five PCR protocols were standardized to assess the presence of genetic material from mycoplasma, porcine circovirus 1 (PCV1), bovine leukemia virus (BLV) or bovine viral diarrhea virus (BVDV) in cell culture samples. PCR reactions for the genes GAPDH and beta-actin were used to evaluate the efficiency of nucleic acid extraction. The PCR protocols were applied to 88 cell culture samples from eight laboratories. The tests were also used to assess potential contamination in 10 trypsin samples and 13 fetal calf serum samples from different lots from five of the laboratories. The results showed the occurrence of the following as DNA cell culture contaminants: 34.1% for mycoplasma, 35.2% for PCV1, 23.9% for BVDV RNA and 2.3% for BLV. In fetal calf sera and trypsin samples BVDV RNA and PCV1 DNA was detected. The results demonstrated that cell culture, sera and trypsin used by different laboratories show a high rate of contaminants. The results highlight the need for monitoring cell cultures and controlling for biological contaminants in laboratories and cell banks working with these materials.
Insights
Polymerase chain reaction (PCR) detected significant contamination in cell cultures, sera, and trypsin. This highlights the critical need for rigorous monitoring of biological contaminants in laboratory materials.
Area of Science:
- Microbiology
- Molecular Biology
- Biotechnology
Background:
- Cell cultures, sera, and trypsin are essential reagents in biological research and biopharmaceutical production.
- Contamination of these materials can compromise experimental results and product safety.
- Standardized methods for contaminant detection are crucial for quality control.
Purpose of the Study:
- To standardize and apply polymerase chain reaction (PCR) techniques for detecting common contaminants in cell cultures, sera, and trypsin.
- To assess the prevalence of specific contaminants including mycoplasma, porcine circovirus 1 (PCV1), bovine leukemia virus (BLV), and bovine viral diarrhea virus (BVDV).
Main Methods:
- Standardization of five PCR protocols for detecting mycoplasma, PCV1, BLV, and BVDV.
- Application of PCR to 88 cell culture samples, 10 trypsin samples, and 13 fetal calf serum samples from multiple laboratories.
- Utilized GAPDH and beta-actin gene PCRs to validate nucleic acid extraction efficiency.
Main Results:
- High contamination rates were observed: 34.1% for mycoplasma, 35.2% for PCV1, 23.9% for BVDV RNA, and 2.3% for BLV in cell cultures.
- Bovine viral diarrhea virus (BVDV) RNA and PCV1 DNA were also detected in fetal calf sera and trypsin samples.
- Significant levels of biological contaminants were found across various laboratories and reagent batches.
Conclusions:
- Cell cultures, sera, and trypsin commonly used in laboratories harbor a high prevalence of contaminants.
- The findings underscore the urgent necessity for routine monitoring and stringent control of biological contaminants in these critical materials.
- Implementing standardized PCR detection methods is vital for ensuring the integrity of research and biopharmaceutical manufacturing.
