Cell culture studies provide a valuable complement to in vivo experiments, allowing for a more controlled manipulation of cellular functions and processes. For decades, cell lines have played a critical role in scientific advancements, yet researchers have become increasingly cautious when interpreting data generated from cell lines only. Factors such as misidentified and contaminated cell lines have spurred renewed interest in primary cells [1, 2]. Moreover, cell lines often differ genetically and phenotypically from their tissue origin, whereas primary cells maintain many of the important markers and functions seen in vivo [3, 4]. Endothelial cell lines, for example, lack various functional markers, while primary endothelial cells retain these critical features. Despite these advantages, obtaining a pure population of primary cells can be a difficult and arduous process. At ScienCell Research Laboratories, where we specialize in primary cell culture, we understand the many challenges that scientists face when working with primary cells.
Primary cells, in contrast to cell lines, are extremely sensitive cells requiring additional nutrients not included in classical media. To optimize survival and growth, primary cells perform best in specialty media customized for each cell type. An endothelial cell, for example, has very different nutritional requirements than an epithelial cell or a neuron, and thus requires a unique medium. Traditional cell culture media has relied on serum to provide the growth factors, hormones, lipids and other undefined components to support cellular growth. For primary cells, however, high serum levels can lead to differentiation or promote growth of contaminating cells like fibroblasts. In addition, serum is plagued by rising costs and lot to lot variability. Formulating specialty media with little or no serum circumvents these issues while enabling greater customization to promote growth of individual primary cell types. Other practices, such as seeding primary cells on more physiologically relevant substrates rather than using synthetic polymers can significantly improve cell attachment, growth, and purity.
Gene expression analysis is critical for understanding the transcriptome profiles of primary cells and how this directly influences their functionality. Traditional reporter-gene assays and cDNA microarrays often require either transfection of exogenous material or large quantities of high quality RNA. Primary cells, however, are notoriously difficult to transfect and the efficiency varies greatly between different cell types. In addition, primary cells have a finite lifespan and limited expansion capacity, making it difficult to obtain a high yield of RNA. To help address these challenges, ScienCell has developed GeneQuery™ qPCR Array kits and custom primer design service for gene expression profiling. GeneQuery™ will enable researchers to directly measure protein expression patterns of primary cells, stem cells, tissues, and cell lines. Importantly, the qPCR arrays have been validated and optimized using primary cell cDNA. Due to the high sensitivity and specificity of the assay, even genes with very low abundance can be analyzed. Gene expression analysis of primary cells will enable researchers to better understand biological pathways and disease processes, such as cell cycle regulation, stem cell biology, cancer development, and neurological disorders.
Primary cells are critical tools in the pursuit of future scientific breakthroughs and therapies. For over fifteen years, ScienCell has worked towards developing the tools necessary for scientists to achieve such advancements.
References:
[1] Alston-Roberts C, et al. (2010) “Cell line misidentification: the beginning of the end.” Nat Rev Cancer. 10(6): 441-448.
[2] Lorsch J, Collins F, Lippincott-Schwartz J. (2014) “Fixing problems with cell lines.” Science. 346 (6216): 1452-1453.
[3] Pan C, Kumar C, Bohl S, Klingmueller U, Mann M. (2009) “Comparative Proteomic Phenotyping of Cell Lines and Primary Cells to Assess Preservation of Cell Type-specific Functions.” Mol Cell Proteomics. 8(3): 443-450.
[4] Alge C, Hauck S, Priglinger S, Kampik A, Ueffing M. (2006) “Differential protein profiling of primary versus immortalized human RPE cells identifies expression patterns associated with cytoskeletal remodeling and cell survival.” J Proteome Res. 5(4): 862-878.
ORIGINALLY Written for and Posted at 2015 Cell Biology ASCB Annual Meeting website
LINK: The Advantages and Difficulties of Working with Primary Cells