The debate over using primary cells versus cell lines continues to challenge researchers. Primary cells are believed to be more biologically relevant tools than cell lines for studying human and animal biology.
With the emergence of newer technologies such as 3D culture, the use of primary cells are becoming increasingly prevalent to achieve improved results.
For decades, cell lines, with their ability to proliferate virtually indefinitely, have been used as a cost-effective tool in basic research. However, some cell lines may not be the cell type they have historically been reported to be. For example, a review about cell line quality summarized that 18–36% of cell lines are misidentified or cross-contaminated (Hughes P et al., 2007).
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Serial passaging of cell lines over time
can cause genotypic and phenotypic variation.
Bioinformatic analysis of proteomic phenotypes
revealed that the Hepa1–6 cell lines were
deficient in mitochondria, reflecting rearrangement
of metabolic pathways when compared
to primary hepatocytes (Pan et al., 2009).
Therefore, researchers should be careful about
only relying on data generated with cell lines.
For example, NIH highlights authentication of
cultured cell lines as critical for grant applications.
And the current recommendation from
the scientific community is to use key control
experiments with primary cells to qualify the
findings seen in cell lines.
|Cell Culture Procedures are Conducted with Two Types of Cells:
||Primary Cells vs. Cell Lines Infographic
Primary Cells – Cells isolated directly from human
or animal tissue using enzymatic or mechanical methods. Once isolated,
they are placed in an artificial environment in plastic or glass
containers supported with specialized medium containing essential
growth factors to support proliferation. Primary cells could be of two
types – adherent or suspension. Adherent cells require attachment for
growth and are said to be anchorage-dependent cells. The adherent cells
are usually derived from tissues of organs. Suspension cells do not
require attachment for growth
and are said to be anchorage-independent cells. All suspension cells are
isolated from blood system. Although primary cells usually have a
limited lifespan, they offer a huge number of advantages compared to
Cell Lines – Cells that have been continually
passaged over a long period of time and have acquired homogenous
genotypic and phenotypic characteristics. Cell lines can be finite or
continuous. An immortalized or continuous cell line
has acquired the ability to proliferate indefinitely, either through
genetic mutations or artificial modifications. A finite cell line has
been sub-cultured for 20-80 passages after which they senesce. Cell
lines are preferably used for
convenience as they are easy to handle and widely published. However,
they are less preferred as a biologically relevant option, since they
have lost the true characteristics of the original tissue from which
they were isolated.
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||Immortalized Cell Lines
||Limited, resembles tissue characteristics
||Infinite, loses tissue characteristics
|Closer to an in vivo model:
||Yes, isolated directly from the tissue
||No, clonally selected over time
|Reduces animal testing costs:
||Yes, used in advanced cell culture models to refine experiments
||Limited ability to develop biologically relevant complex in vitro models
|Authentication required before use:
||No, if bought commercially
||Yes, mandated by many government institutes and scientific journals
|Availability of donor characteristics:
Primary Cells and Cell Lines Show Variability in
Drug Dose Response
Figure 1. Primary cells and cell lines show variability in drug dose, so data acquired through cell lines cannot easily be replicated in an in vivo model. The three cell types were dosed with Camptothecin for 48h and their cytotoxic response assessed using the ViaLight™ Assay.The ViaLight™ Assay measures the number of viable cells left after drug challenge, measuring the ATP donated to the assay reaction by viable cells. In this assay the data points are the mean of 40 replicates (8 replicates per plate; 5 plates). We have a marked difference in EC50 dose response almost 10 fold. The HEK 293 cell line required only 68nM Camptothecin to kill 50% of the cells. In the case of the primary cells (HRE = human renal epithelial cells, HRCE = human renal cortical epithelial cells) this dose was over 500nM a strikingly different response.
Figure 2. Some pathways can only be studied in primary cells, as there is no cell line to represent that pathway. Cytolytic T lymphocytes (CTLs) recognize foreign cells and lyse them via the “immunological synapse”. At the immunological synapse CTLs secrete cytolytic granules containing Perforin. hMUNC13-4 is involved in an activation step preceding vesicle membrane fusion in cytolytic granule secretion. Nucleofection™of a plasmid expressing hMUNC13-4/RFP fusion shows only partial co-localization of hMUNC13-4 (red, H) and Perforin (green, I) in isolated CTLs (overlay J; blue: nuclei stained by DAPI). Upon target cell contact (M) hMUNC13-4 polarizes and extensively co-localizes with Perforincontaining cytolytic granules at the immunological synapse (L). Reprinted from: Feldmann J et al. (2003) Cell 115(4):461-73; with permission of Elsevier.
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Some Pathways Can Only Be Studied in Primary Cells