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Mononuclear cells contaminating acute lymphoblastic leukaemic samples tested for cellular drug resistance using the methyl-thiazol-tetrazolium assay.

The methyl-thiazol-tetrazolium (MTT) assay is a drug resistance assay which cannot discriminate between malignant and non-malignant cells. We previously reported that samples with > or = 80% leukaemic cells at the start of culture give similar results in the MTT assay and the differential stainin...

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Autores principales: Kaspers, G. J., Veerman, A. J., Pieters, R., Broekema, G. J., Huismans, D. R., Kazemier, K. M., Loonen, A. H., Rottier, M. A., van Zantwijk, C. H., Hählen, K.
Formato: Texto
Lenguaje:English
Publicado: Nature Publishing Group 1994
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2033662/
https://www.ncbi.nlm.nih.gov/pubmed/7981053
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author Kaspers, G. J.
Veerman, A. J.
Pieters, R.
Broekema, G. J.
Huismans, D. R.
Kazemier, K. M.
Loonen, A. H.
Rottier, M. A.
van Zantwijk, C. H.
Hählen, K.
author_facet Kaspers, G. J.
Veerman, A. J.
Pieters, R.
Broekema, G. J.
Huismans, D. R.
Kazemier, K. M.
Loonen, A. H.
Rottier, M. A.
van Zantwijk, C. H.
Hählen, K.
author_sort Kaspers, G. J.
collection PubMed
description The methyl-thiazol-tetrazolium (MTT) assay is a drug resistance assay which cannot discriminate between malignant and non-malignant cells. We previously reported that samples with > or = 80% leukaemic cells at the start of culture give similar results in the MTT assay and the differential staining cytotoxicity assay, in which a discrimination between malignant and non-malignant cells can be made. However, the percentage of leukaemic cells may change during culture, which might affect the results of the MTT assay. We studied 106 untreated childhood acute lymphoblastic leukemia (ALL) samples with > or = 80% leukaemic cells at the start of culture. This percentage decreased below 80% in 28%, and below 70% in 13%, of the samples after 4 days of culture. A decrease below 70% occurred more often in case of 80-89% leukaemic cells (9/29) than in case of > or = 90% leukaemic cells at the start of culture (5/77, P = 0.0009). Samples with < 70% leukaemic cells after culture were significantly more resistant to 6 out of 13 drugs, and showed a trend towards being more resistant to two more drugs, than samples with > or = 80% leukaemic cells. No such differences were seen between samples with 70-79% and samples with > or = 80% leukaemic cells after culture. We next studied in another 30 ALL samples whether contaminating mononuclear cells could be removed by using immunoamagnetic beads. Using a beads to target cell ratio of 10:1, the percentage of leukaemic cells increased from mean 72% (s.d. 9.3%) to mean 87% (s.d. 6.7%), with an absolute increase of 2-35%. The recovery of leukaemic cells was mean 82.1% (range 56-100%, s.d. 14.0%). The procedure itself did not influence the results of the MTT assay in three samples containing only leukaemic cells. We conclude that it is important to determine the percentage of leukaemic cells at the start and at the end of the MTT assay and similar drug resistance assays. Contaminating mononuclear cells can be successfully removed from ALL samples using immunomagnetic beads. This approach may increase the number of leukaemic samples which can be evaluated for cellular drug resistance with the MTT assay or a similar cell culture drug resistance assay.
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spelling pubmed-20336622009-09-10 Mononuclear cells contaminating acute lymphoblastic leukaemic samples tested for cellular drug resistance using the methyl-thiazol-tetrazolium assay. Kaspers, G. J. Veerman, A. J. Pieters, R. Broekema, G. J. Huismans, D. R. Kazemier, K. M. Loonen, A. H. Rottier, M. A. van Zantwijk, C. H. Hählen, K. Br J Cancer Research Article The methyl-thiazol-tetrazolium (MTT) assay is a drug resistance assay which cannot discriminate between malignant and non-malignant cells. We previously reported that samples with > or = 80% leukaemic cells at the start of culture give similar results in the MTT assay and the differential staining cytotoxicity assay, in which a discrimination between malignant and non-malignant cells can be made. However, the percentage of leukaemic cells may change during culture, which might affect the results of the MTT assay. We studied 106 untreated childhood acute lymphoblastic leukemia (ALL) samples with > or = 80% leukaemic cells at the start of culture. This percentage decreased below 80% in 28%, and below 70% in 13%, of the samples after 4 days of culture. A decrease below 70% occurred more often in case of 80-89% leukaemic cells (9/29) than in case of > or = 90% leukaemic cells at the start of culture (5/77, P = 0.0009). Samples with < 70% leukaemic cells after culture were significantly more resistant to 6 out of 13 drugs, and showed a trend towards being more resistant to two more drugs, than samples with > or = 80% leukaemic cells. No such differences were seen between samples with 70-79% and samples with > or = 80% leukaemic cells after culture. We next studied in another 30 ALL samples whether contaminating mononuclear cells could be removed by using immunoamagnetic beads. Using a beads to target cell ratio of 10:1, the percentage of leukaemic cells increased from mean 72% (s.d. 9.3%) to mean 87% (s.d. 6.7%), with an absolute increase of 2-35%. The recovery of leukaemic cells was mean 82.1% (range 56-100%, s.d. 14.0%). The procedure itself did not influence the results of the MTT assay in three samples containing only leukaemic cells. We conclude that it is important to determine the percentage of leukaemic cells at the start and at the end of the MTT assay and similar drug resistance assays. Contaminating mononuclear cells can be successfully removed from ALL samples using immunomagnetic beads. This approach may increase the number of leukaemic samples which can be evaluated for cellular drug resistance with the MTT assay or a similar cell culture drug resistance assay. Nature Publishing Group 1994-12 /pmc/articles/PMC2033662/ /pubmed/7981053 Text en https://creativecommons.org/licenses/by/4.0/This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit https://creativecommons.org/licenses/by/4.0/.
spellingShingle Research Article
Kaspers, G. J.
Veerman, A. J.
Pieters, R.
Broekema, G. J.
Huismans, D. R.
Kazemier, K. M.
Loonen, A. H.
Rottier, M. A.
van Zantwijk, C. H.
Hählen, K.
Mononuclear cells contaminating acute lymphoblastic leukaemic samples tested for cellular drug resistance using the methyl-thiazol-tetrazolium assay.
title Mononuclear cells contaminating acute lymphoblastic leukaemic samples tested for cellular drug resistance using the methyl-thiazol-tetrazolium assay.
title_full Mononuclear cells contaminating acute lymphoblastic leukaemic samples tested for cellular drug resistance using the methyl-thiazol-tetrazolium assay.
title_fullStr Mononuclear cells contaminating acute lymphoblastic leukaemic samples tested for cellular drug resistance using the methyl-thiazol-tetrazolium assay.
title_full_unstemmed Mononuclear cells contaminating acute lymphoblastic leukaemic samples tested for cellular drug resistance using the methyl-thiazol-tetrazolium assay.
title_short Mononuclear cells contaminating acute lymphoblastic leukaemic samples tested for cellular drug resistance using the methyl-thiazol-tetrazolium assay.
title_sort mononuclear cells contaminating acute lymphoblastic leukaemic samples tested for cellular drug resistance using the methyl-thiazol-tetrazolium assay.
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2033662/
https://www.ncbi.nlm.nih.gov/pubmed/7981053
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