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Standard melanoma-associated markers do not identify the MM127 metastatic melanoma cell line.

Haridas P, McGovern JA, Kashyap AS, McElwain DL, Simpson MJ - Sci Rep (2016)

Bottom Line: Common markers used to identify melanoma cell lines include: S100; HMB-45; and Melan-A.Our results show that the metastatic cell line, MM127, cannot be detected using any of the commonly used melanoma-associated markers.This implies that it would be very difficult to identify this particular cell line in a heterogeneous sample, and as a result this cell line should be used with care.

View Article: PubMed Central - PubMed

Affiliation: Institute of Health and Biomedical Innovation, Queensland University of Technology (QUT), Kelvin Grove 4059, Australia.

ABSTRACT
Reliable identification of different melanoma cell lines is important for many aspects of melanoma research. Common markers used to identify melanoma cell lines include: S100; HMB-45; and Melan-A. We explore the expression of these three markers in four different melanoma cell lines: WM35; WM793; SK-MEL-28; and MM127. The expression of these markers is examined at both the mRNA and protein level. Our results show that the metastatic cell line, MM127, cannot be detected using any of the commonly used melanoma-associated markers. This implies that it would be very difficult to identify this particular cell line in a heterogeneous sample, and as a result this cell line should be used with care.

No MeSH data available.


Related in: MedlinePlus

MM127 cell line does not express three standard melanoma-associated markers.(a) STR analysis of the MM127 cell line. Upper row shows the allele names. Lower row describes the location of the allele on the chromosome. (b) Immunofluorescence results for four melanoma cell lines (WM35, WM793, SK-MEL-28, MM127). Cells are fixed in 4% paraformaldehyde and stained for S100 (green), HMB-45 (green) and Melan-A (green). The nucleus (blue) and f-actin (red) are highlighted. Results for the negative control (HaCaT) are given in the supplementary material [Fig. S1]. (c) Melanoma (WM35, WM793, SK-MEL-28 and MM127) and negative controls (primary cells: keratinocytes, fibroblasts, and cell line: HaCaT) are analysed by Western blotting for S100, HMB-45 and Melan-A. GAPDH is used as loading control and detected at 37 kDa. (d) qRT-PCR results show the difference between the expression of melanoma-associated genes for the negative control (HaCaT) and various cell lines. Values correspond to the mean ΔCt (n = 3), where ΔCt = Ct (RPL32) − Ct (target gene). Error bars indicate the standard error (n = 3). Results below the detectable limit are given as BDL.
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f1: MM127 cell line does not express three standard melanoma-associated markers.(a) STR analysis of the MM127 cell line. Upper row shows the allele names. Lower row describes the location of the allele on the chromosome. (b) Immunofluorescence results for four melanoma cell lines (WM35, WM793, SK-MEL-28, MM127). Cells are fixed in 4% paraformaldehyde and stained for S100 (green), HMB-45 (green) and Melan-A (green). The nucleus (blue) and f-actin (red) are highlighted. Results for the negative control (HaCaT) are given in the supplementary material [Fig. S1]. (c) Melanoma (WM35, WM793, SK-MEL-28 and MM127) and negative controls (primary cells: keratinocytes, fibroblasts, and cell line: HaCaT) are analysed by Western blotting for S100, HMB-45 and Melan-A. GAPDH is used as loading control and detected at 37 kDa. (d) qRT-PCR results show the difference between the expression of melanoma-associated genes for the negative control (HaCaT) and various cell lines. Values correspond to the mean ΔCt (n = 3), where ΔCt = Ct (RPL32) − Ct (target gene). Error bars indicate the standard error (n = 3). Results below the detectable limit are given as BDL.

Mentions: The alleles obtained from STR profiling are analysed using the DMSZ database (http://www.dsmz.de/fp/cgi-bin/str.html) to give the closest match to each cell line we consider. The results from this analysis confirm that WM35, WM793 and SK-MEL-28 are as expected (Supplementary information). Conversely, the results for MM127 are not as expected since there is no match identified using the MM127 alleles [Fig. 1(a)]. This preliminary result suggests that further investigation is warranted.


Standard melanoma-associated markers do not identify the MM127 metastatic melanoma cell line.

Haridas P, McGovern JA, Kashyap AS, McElwain DL, Simpson MJ - Sci Rep (2016)

MM127 cell line does not express three standard melanoma-associated markers.(a) STR analysis of the MM127 cell line. Upper row shows the allele names. Lower row describes the location of the allele on the chromosome. (b) Immunofluorescence results for four melanoma cell lines (WM35, WM793, SK-MEL-28, MM127). Cells are fixed in 4% paraformaldehyde and stained for S100 (green), HMB-45 (green) and Melan-A (green). The nucleus (blue) and f-actin (red) are highlighted. Results for the negative control (HaCaT) are given in the supplementary material [Fig. S1]. (c) Melanoma (WM35, WM793, SK-MEL-28 and MM127) and negative controls (primary cells: keratinocytes, fibroblasts, and cell line: HaCaT) are analysed by Western blotting for S100, HMB-45 and Melan-A. GAPDH is used as loading control and detected at 37 kDa. (d) qRT-PCR results show the difference between the expression of melanoma-associated genes for the negative control (HaCaT) and various cell lines. Values correspond to the mean ΔCt (n = 3), where ΔCt = Ct (RPL32) − Ct (target gene). Error bars indicate the standard error (n = 3). Results below the detectable limit are given as BDL.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4834532&req=5

f1: MM127 cell line does not express three standard melanoma-associated markers.(a) STR analysis of the MM127 cell line. Upper row shows the allele names. Lower row describes the location of the allele on the chromosome. (b) Immunofluorescence results for four melanoma cell lines (WM35, WM793, SK-MEL-28, MM127). Cells are fixed in 4% paraformaldehyde and stained for S100 (green), HMB-45 (green) and Melan-A (green). The nucleus (blue) and f-actin (red) are highlighted. Results for the negative control (HaCaT) are given in the supplementary material [Fig. S1]. (c) Melanoma (WM35, WM793, SK-MEL-28 and MM127) and negative controls (primary cells: keratinocytes, fibroblasts, and cell line: HaCaT) are analysed by Western blotting for S100, HMB-45 and Melan-A. GAPDH is used as loading control and detected at 37 kDa. (d) qRT-PCR results show the difference between the expression of melanoma-associated genes for the negative control (HaCaT) and various cell lines. Values correspond to the mean ΔCt (n = 3), where ΔCt = Ct (RPL32) − Ct (target gene). Error bars indicate the standard error (n = 3). Results below the detectable limit are given as BDL.
Mentions: The alleles obtained from STR profiling are analysed using the DMSZ database (http://www.dsmz.de/fp/cgi-bin/str.html) to give the closest match to each cell line we consider. The results from this analysis confirm that WM35, WM793 and SK-MEL-28 are as expected (Supplementary information). Conversely, the results for MM127 are not as expected since there is no match identified using the MM127 alleles [Fig. 1(a)]. This preliminary result suggests that further investigation is warranted.

Bottom Line: Common markers used to identify melanoma cell lines include: S100; HMB-45; and Melan-A.Our results show that the metastatic cell line, MM127, cannot be detected using any of the commonly used melanoma-associated markers.This implies that it would be very difficult to identify this particular cell line in a heterogeneous sample, and as a result this cell line should be used with care.

View Article: PubMed Central - PubMed

Affiliation: Institute of Health and Biomedical Innovation, Queensland University of Technology (QUT), Kelvin Grove 4059, Australia.

ABSTRACT
Reliable identification of different melanoma cell lines is important for many aspects of melanoma research. Common markers used to identify melanoma cell lines include: S100; HMB-45; and Melan-A. We explore the expression of these three markers in four different melanoma cell lines: WM35; WM793; SK-MEL-28; and MM127. The expression of these markers is examined at both the mRNA and protein level. Our results show that the metastatic cell line, MM127, cannot be detected using any of the commonly used melanoma-associated markers. This implies that it would be very difficult to identify this particular cell line in a heterogeneous sample, and as a result this cell line should be used with care.

No MeSH data available.


Related in: MedlinePlus