A comparison of flow cytometry and immunohistochemistry in human colorectal cancers

Diederichsen, Axel Cosmus Pyndt; Hansen, Tine Plato; Nielsen, Ole; Fenger, Claus; Jensenius, Jens Christian; Christensen, P.B.; Kristensen, Tom; Zeuthen, Jesper

doi:10.1111/j.1699-0463.1998.tb01385.x

Cited by 14 publications

(15 citation statements)

References 21 publications

(11 reference statements)

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“…Using flow cytometry, we could not distinguish between tumour cells and normal epithelial cells (both of which are EP4 positive). However we have observed that the majority, if not all, of the EP4 positive cells represents tumour cells (Diederichsen et al, 1997) and these are counted as such. We found that HLA class I was expressed on all tumour cells in 60 of the tumours, and reduced in the remainder 10 (15-99% of the tumour cells expressed HLA class I).…”

Section: Discussionmentioning

confidence: 94%

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Flow cytometric investigation of immune-response-related surface molecules on human colorectal cancers

et al. 1998

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Our purpose was to clarify whether human colorectal cancer cells are equipped to present tumour‐associated‐antigens to the immune system, and whether this ability correlates with lymphoid infiltration, the Dukes' stage and Jass classification. Enzymatically dissociated tumour cells from 70 different colorectal cancers were monitored by multiparameter flow cytometry. Gating on EP4+ cells, the expression of the surface molecules HLA class I, HLA class II, CD80 (B7‐1), CD54 (ICAM‐1) and CD58 (LFA‐3) was evaluated. In 60 of 70 tumours, all tumour cells expressed HLA class I, in 10 tumours 15‐96% of the tumour cells expressed HLA class I. In 1 tumour, all tumour cells expressed HLA class II, in 67 tumours some expressed HLA class II, in 2 tumours none expressed HLA class II. Expression of CD58 was heterogeneous, and there was no or only sparse expression of CD80 and CD54. Expression of the HLA class I molecules, but not the class II, was correlated with lymphoid infiltration and the Jass classification. Expression of these surface molecules was not correlated with the Dukes' stage. The tumour cells were generally equipped to present antigens to the effector arm of the immune system since HLA class I is expressed, but the tumour cells were not optimal in stimulating an immune response, since HLA class II and CD58 were only marginally expressed and CD80 and CD54 were absent. Int. J. Cancer (Pred. Oncol.) 79:283–287, 1998.© 1998 Wiley‐Liss, Inc.

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Section: Discussionmentioning

confidence: 94%

“…These data were obtained by immunohistochemical analysis. Flow cytometry has been shown to be more sensitive (Coventry et al, 1994), and tumours previously described as HLA class I negative by immunohistochemistry were found to express HLA class I by flow cytometry (Diederichsen et al, 1997).…”

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confidence: 99%

Flow cytometric investigation of immune-response-related surface molecules on human colorectal cancers

et al. 1998

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“…IHC has also been the method of choice to assess the presence of tumor cells within TIL cultures for adoptive transfer therapy. However, IHC is labor intensive, requires large number of slides, does not allow an assessment of viability, is technically challenging, is semi-quantitative and requires a trained eye and/or pathologist (11)(12)(13). The amount of time involved in performing IHC has decreased by utilizing automated microscopy; however, manual review is still required to confirm positive cells (12).…”

Section: Discussionmentioning

confidence: 99%

“…Second, IHC and PCR are unable to distinguish viable cells from dead cells, where as impermeant fluorescent dyes such as 7-AAD or the amine-reactive dyes have increased fluorescent signals in dead cells that are easily distinguished by flow cytometry. Third, flow cytometry may be superior to IHC in the detection of weakly expressed antigens (11). Finally, IHC is semi-quantitative (11) where an estimate is given of the percent positive.…”

Section: Discussionmentioning

confidence: 99%

“…However, there are several disadvantages to IHC. The assay is unable to detect viable cells, it is subjective in interpretation, and it semiquantitative at best (11,12). Advantages of flow cytometry include its ability to acquire multiple parameters and large numbers of data points in a short period of time which make it an ideal technology to evaluate cellular purity in a qualitative and quantitative manner.…”

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Flow cytometry assessment of residual melanoma cells in tumor‐infiltrating lymphocyte cultures

et al. 2012

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Adoptive transfer of tumor-infiltrating lymphocytes (TIL) is in development for the treatment of metastatic melanoma. In phase II clinical trials, patients with metastatic melanoma that received TIL after preconditioning had a 50-70% clinical response rate. The current approach to generate TIL is to culture melanoma enzyme digests in the presence of IL-2 for a 10-to 20-day period followed by 2 weeks of rapid expansion (REP). Prior to administration, cell therapies are characterized and tested for purity. TIL are characterized by CD3 surface marker expression, and purity is assessed by the amount of tumor remaining in culture. Evaluating TIL purity has traditionally been done by immunohistochemistry, which is often considered semiquantitative. To generate a quantitative assay, we used multiparameter flow cytometry to evaluate the presence of viable tumor cells by staining TIL populations with a viability dye and an antibody cocktail that detects intracellular tumor-antigens gp100, Mart-1, tyrosinase, S100, and surface tumor-antigen melanoma chondroitin sulfate proteoglycan (MCSP), and CD3 on T cells. Tumors were identified by gating on the viable CD3 2 population. Antigens in tumors were initially optimized with individual antibodies using both immunohistochemistry and flow cytometry. When eight different tumor cell lines were spiked into an activated T cell culture, flow cytometry was able to distinguish lymphocytes from tumors in all samples tested. Most importantly, the assay was able to detect melanoma cells in all enzyme digests (9/9) from patient samples. After IL-2-induced TIL expansion, there was a significant decrease in tumor cells; tumor cells were detected in only 2 of 12 samples. In eight IL-2-induced TIL samples that were further expanded in REP, no tumor cells were detected. We have demonstrated that flow cytometry is an alternative to immunohistochemistry for defining the purity of a TIL population. ' 2012 International Society for Advancement of Cytometry

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HLA‐DR expression is associated with better prognosis in sporadic Australian clinicopathological Stage C colorectal cancers

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et al. 2009

Intl Journal of Cancer

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Predicting patient outcome for colorectal carcinoma (CRC) with lymph node but not distant metastases remains challenging. Various prognostic markers have been identified including microsatellite instability (MSI) and possibly expression of the MHC Class II protein, HLA-DR. About 15% of sporadic CRC exhibits MSI associated with methylation of the DNA mismatch repair gene hMLH1 promoter. In addition, a significant proportion of unselected CRC demonstrates expression of HLA-DR. We sought to examine the relationship between HLA-DR expression, MSI status and prognosis in sporadic Australian Clinicopathological (ACP) Stage C CRC. Two hundred seventy consecutive patients with sporadic ACP Stage C CRC were treated at Concord Repatriation General Hospital between 1986 and 1992. None of these patients received adjuvant chemotherapy and all were followed for a minimum of 5 years or until death. DNA was extracted from paraffin sections and MSI status determined by PCR. HLA-DR expression was determined immunohistochemically using an antibody against the HLA-DR a chain. MSI status could be assigned in 235 cases: 176 CRCs (74.9%) were microsatellite stable, whereas 23 (9.8%) had high levels of MSI (MSI-H) and 36 (15.3%) had low levels of MSI (MSI-L). HLA-DR expression by CRC cells was seen in 148 (60.1%) cases and correlated with the presence of tumor-infiltrating lymphocytes (p 5 0.0005) and peritumoral lymphocytes (p 5 0.003), but not other clinicopathological features or MSI status. HLA-DR-positive CRCs were strongly associated with better patient outcome (p < 0.0001). ' 2009 UICC

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A comparison of flow cytometry and immunohistochemistry in human colorectal cancers

Cited by 14 publications

References 21 publications

Flow cytometric investigation of immune-response-related surface molecules on human colorectal cancers

Flow cytometric investigation of immune-response-related surface molecules on human colorectal cancers

Flow cytometry assessment of residual melanoma cells in tumor‐infiltrating lymphocyte cultures

HLA‐DR expression is associated with better prognosis in sporadic Australian clinicopathological Stage C colorectal cancers

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