Host Cell DNA (HCD) Detection and Analysis Service

Residual DNA (rDNA) refers to the aggregate of deoxyribonucleic acid (DNA) and fragments extracted from recombinant host cells, commonly termed as  host cell residual DNA (HCD). Currently, there are various types of animal cell substrates used for the production of vaccines and other biologics, such as primary cells, passaged cells, and human diploid cells. Primary cells include hamster kidney cells, guinea pig kidney cells, bovine kidney cells, monkey kidney cells, and chicken embryo fibroblasts, which are mainly used for the production of live attenuated vaccines and inactivated vaccines. Passaged cells refer to cell lines that can divide and propagate indefinitely in culture. Commonly used passaged cells include Madin-Darby canine kidney (MDCK) cells and African green monkey kidney (Vero) cells.

 

Associated risks with rDNA span infectivity (via viruses like HIV), carcinogenicity (through oncogenes such as Ras), immunogenicity (via CpG-rich sequences in bacteria), and mutagenicity (through mechanisms like transposons, retrotransposons, and DNA recombination). In 1987, WHO guidelines advised that total rDNA should not surpass 100 pg per dose, which was subsequently adjusted to 10 ng per dose. The European Pharmacopoeia generally limits residual DNA in bioproducts to no more than 10 ng per dose, with stricter standards for certain vaccines; for instance, the DNA residue in inactivated hepatitis A vaccines must not exceed 100 pg per dose, and in hepatitis B vaccines, no more than 10 pg per dose. The inherent risks of rDNA in products designed for human use, coupled with the regulatory mandates to verify its elimination in bioprocessing, underscore the importance of stringent monitoring of rDNA content in the final pharmaceutical product. Detection methodologies must be precise, sensitive, and quantitative to affirm that the DNA clearance meets established standards. Over the course of biotechnological product development, the analytical techniques for rDNA evaluation have evolved continually.

 

MtoZ Biolabs boasts a comprehensive platform for host cell nucleic acid (HCD) and HCR) analysis, along with a development platform for related nucleic acid testing products. The services offered by our platform include the identification of various HCD species, identification of various HCR species, analysis of HCD fragment length and distribution, and the development of related HCD/HCR assay kits. This enables research in areas such as nucleic acid extraction, quantitative nucleic acid testing, and fragment length analysis. Our team employs multi-gradient experiments to ascertain optimal experimental conditions, utilizes methods such as real-time fluorescence quantitative PCR and capillary electrophoresis for data collection, and applies bioinformatics to quantify and identify HCD/HCR. MtoZ Biolabs provides a suite of services including extraction, quantification, and analysis of HCD/HCR, alongside kit development, furnishing a comprehensive solution for analyzing host cell nucleic acids.

• • HCD Fragment Detection Service With CE and qPCR

  Apart from the quantity of DNA, the size distribution of residual DNA fragments is also an important factor related to carcinogenicity and infectivity. Due to the low content of residual DNA in drugs, the development of residual DNA size detection with high sensitivity, specificity, and resolution is a significant challenge.

• • Comprehensive Host Cell DNA (HCD) Detection Service

  Several methodologies are employed for quantifying rDNA within biological processes. These include traditional methods (PicoGreen method, hybridization method, and threshold method) as well as more commonly used methods nowadays such as quantitative real-time PCR (qPCR), capillary electrophoresis (CE), and primer-based whole genome amplification (pWGA) technology.

• • Comprehensive DNA Extraction and Purification Service

  The sample preparation process encompasses: (1) releasing DNA from samples and cells through lysis, (2) isolating DNA, (3) purifying DNA by removing inhibitory substances, and (4) optionally, concentrating DNA, which is critical for detecting low-concentration analytes.

• • Comprehensive Host Cell Residual RNA Detection Service

  DNA products such as plasmid/minicircle DNA can serve as primary part of DNA vaccine products or as important raw material for viral vectors or DNA vector of gene therapy, and for viral vector or non-viral vector of cellular immunotherapy. Residual RNA in these DNA products may affect  biological activity of these products (such as interfering with plasmid transfection and expression efficiency) or pose risks (such as triggering immune responses by activating the cellular interferon pathway due to exogenous