Identification and verification of rodent cell lines by polymerase chain reaction

Abstract

Cell lines represent valuable tools for basic research and diagnostic applications as well as for the production of biological products such as antibodies or vaccines. For all cell culturists, a well-identified origin of their cell lines as well as the periodic re-examination of their identity should be a basic requirement. We established a simple polymerase chain reaction (PCR) to verify or identify rodent and human cell lines. Since mouse-, rat-, Chinese hamster- and Syrian hamster-derived cell lines represent the most frequently used rodent cell lines, our investigations were focused on these species. Our assay used oligonucleotide primers annealing to sequences within the beta-actin and the beta-globin gene and to repetitive DNA. Primers were designed mostly from intron sequences of the genes aiming to amplify only one specific DNA segment and thus enabling to exclude easily false DNA. More than 130 cells lines originating from the five species were analyzed in that study. Our PCR revealed specific profiles for all species investigated. No further methods like DNA sequencing or fragment length polymorphism analysis were needed to differentiate these species. The results introduce our PCR-assay as a rapid, specific and routinely feasible tool in order to identify or distinguish rodent cell lines from each other and from human cell lines.

The necessity of identity assessment of animal intestinal cell lines: A case report

“…During the last years scientists at the DSMZ and other institutions have established easy-to-use methods for the analyses of animal cell lines (Stacey et al 1997;Liu et al 2003;Cooper et al 2007;Ono et al 2007;Steube et al 2008). However, many cell lines, ) and waste of time and money due to invalid data can only be estimated (Stacey et al 2000;Chatterjee 2007).…”

“…PCR amplification and electrophoretic analysis of the products were described recently (Steube et al 2003(Steube et al , 2008. Genomic sequencing of the PCR products was conducted by Eurofins Table 2 lists the applied primers for the identification of the animal species.…”

Species identification and authentification of human and rodent cell cultures using polymerase chain reaction analysis of vomeronasal receptor genes

“…Inter-species culture contamination is a common occurrence with the most frequently used cell lines being of human, mouse and rat origin (Steube et al 2008). While assays based on karyotyping, fluorescent in situ hybridization and isozyme analysis (Nardone 2007) have been developed to distinguish between cell lines from different species their application and interpretation can require significant expertise and costs which can inhibit their uptake.…”

“…Yet despite the burgeoning information present in the genome databases for these species only a handful of PCR-based technologies for cell line authentification have been developed (Stacey et al 1997;Parodi et al 2002;Liu et al 2003;López-Andreo et al 2005;Steube et al 2003Steube et al , 2008. Whilst these approaches may be relatively inexpensive many of these assays utilise multiple primers or require interpretation of relatively complex gel banding patterns.…”

Isolation and Propagation of Circulating Tumor Cells from a Mouse Cancer Model

“…Perform a polymerase chain reaction (PCR)-based method, to amplify only one specific DNA segment of the mouse β-globin gene to confirm that the novel established CTC cell lines are mouse cells like the original implanted BNL 1ME A.7R.1 HCC line. Method was originally described and validated by Steube et al 5,8 2. Perform immunostaining for the hepatocyte-specific marker cAMP responsive element binding protein 3-like 3 (CREB3L3) by using a specific antibody.…”