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Bibliography

2011

Human Blood Stabilisation - Immunophenotyping

KEY PAPER: CD4 Intragenic SNPs Associate With HIV-2 Plasma Viral Load and CD4 Count in a Community-Based Study From Guinea-Bissau, West Africa

Sample Type: Lymphocyte subset

Analysis Method: Flow Cytometry

Authors

Branwen Hennig , Digna Velez-Edwards, Maarten Schim van der Loeff, Cyrille Bisseye, Todd Edwards, Alessandra Tacconelli, Giuseppe Novelli, Peter Aaby, Steve Kaye, William Scott, Assan Jaye, Hilton Whittle, Scott Williams, Adrian Hill, Giorgio Sirugo

Publication

Journal of Acquired Immune Deficiency Syndromes (2011) 56, 1-8

A second, back-up method for lymphocyte subset measuring was also in place. For this method, 100 μl "TransFix" solution (Cytomark UK) 24 was added to 500 μl fresh blood and roller-mixed for 5 minutes, after which the sample was kept in a refrigerator at 4 degrees Celsius and transported to the MRC Laboratories in Fajara, The Gambia. When the routine method failed, the transfixed samples were stained with fluorochrome-conjugated monoclonal antibodies manually rather than by Q-Prep, and analyzed in the FACS calibur, using the MultiSet software (Becton Dickinson, Erembodegem, Belgium). Results were regarded as invalid if the CD3 % was < 45%, or if the sum of CD4 % and CD8 % was more than 10% different from the mean CD3 %. In order to investigate whether the results of the two methods (routine and back-up) were comparable, 13 samples were processed using both methods and results compared. The agreement was excellent (r2 = 0.89 for CD4 %). In addition, a Bland and Altman test 25 was performed to compare the two methods and indicated no difference between them (p = 0.3).

http://www.ncbi.nlm.nih.gov/pubmed/20924289

Bone Marrow Stabilisation

KEY PAPER: Immunophenotypic Characterization of Bone Marrow Mast Cells in Mastocytosis and Other Mast Cell Disorders

Sample Type: Bone marrow (mast cells)

Analysis Method: Flow Cytometry

Authors

L. Sánchez-Munoz, C, Teodósio, J.M. Morgado and L. Escribano

Publication

Recent Advances in Cytometry, Part B - Methods in Cell Biology (Book Series), Vol. 103. P333-359, Book published by Academic Press June 2011

Mastocytosis is a term used to designate a heterogeneous group of disorders characterized by an abnormal proliferation and accumulation of mast cells (MCs) in one or multiple tissues including skin, bone marrow (BM), liver, spleen, and lymph nodes, among others. Recent advances in our understanding of mast cell biology and disease resulted in the identification of important differences in the expression of mast cell surface antigens between normal and neoplastic mast cells. Most notably, detection of aberrant expression of CD25 and CD2 on the surface of neoplastic mast cells but not on their normal counterparts lead to the inclusion of this immunophenotypic abnormality in the World Health Organization diagnostic criteria for systemic mastocytosis. Aberrant mast cell surface marker expression can be detected in the bone marrow aspirate by flow cytometry, even in patients lacking histopathologically detectable aggregates of mast cells in bone marrow biopsy sections. These aberrant immunophenotypic features are of great relevance for the assessment of tissue involvement in mastocytosis with consequences in the diagnosis, classification, and follow-up of the disease and in its differential diagnosis with other entities. In this chapter, we provide the reader with information for the objective and reproducible identification of pathologic MCs by using quantitative multiparametric flow cytometry, for their phenotypic characterization, and the criteria currently used for correct interpretation of the immunophenotypic results obtained. In cases in which it is not expected to perform sample processing within the first 24h following a bone marrow puncture, a stabilising solution such as TransFix should be used to avoid deterioration of cells.

http://www.ncbi.nlm.nih.gov/pubmed/21722810


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