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External quality assessment of flow cytometric HLA-B27 typing. 2000

W H Levering, and R van den Beemd, and J G te Marvelde, and W A van Beers, and H Hooijkaas, and K Sintnicolaas, and J W Gratama
Laboratory for Histocompatibility and Immunogenetics, Blood Bank Rotterdam, The Netherlands. bloodrtd@wxs.nl

A biannual external quality assurance (EQA) scheme for flow cytometric typing of the HLA-B27 antigen is operational in The Netherlands and Belgium since 1995. We report here on the results of the first seven send-outs to which 36 to 47 laboratories participated. With the send-out, four specimens from blood bank donors, who had been typed for HLA Class I antigens by complement-dependent cytotoxicity, were distributed. Subtyping of the HLA-B27 allele was performed by PCR-SSP. Ten samples were HLA-B27(pos) (all HLA-B*2705) and 18 were HLA-B27(neg). For flow cytometry, the most widely monoclonal antibody (MoAb) used was FD705, followed by GS145.2 and ABC-m3. The majority of laboratories used more than 1 anti-HLA-B27 MoAb for typing. The HLA-B27(pos) samples were correctly classified as positive by the large majority of participants (median 95%; range 85% to 100% per send out); some participants considered further typing necessary and misclassification as negative was only sporadically seen. The classification of HLA-B27(neg) samples as negative was less straightforward. Ten samples were correctly classified as such by 97% (82% to 100%) of the participants, whereas 64% (range 53% to 70%) of the participants classified the remaining eight samples as HLA-B27(neg). There was no significant prevalence of a particular HLA-B allele among these eight "poor concordancy" samples as compared to the ten "good concordancy" samples. Inspection of the reactivity patterns of the individual MoAb with HLA-B27(neg) samples revealed that ABC-m3 showed very little cross-reactivity apart from its well-known cross-reactivity with HLA-B7, whereas the cross-reactivity patterns of GS145.2 and FD705 were more extensive. The small sample size (n = 18) and the distribution of HLA-B alleles other than HLA-B27 did not allow assignment of specificities to these cross-reactions. Finally, we showed that standardized interpretation of the combined results of two anti-HLA-B27 MoAb reduced the frequency of false-positive conclusions on HLA-B27(neg) samples. In this series, the lowest frequency of false-positive assignments was observed with the combination of the FD705 and ABC-m3 MoAb.

UI MeSH Term Description Entries
D007755 Laboratories, Hospital Hospital facilities equipped to carry out investigative procedures. Hospital Laboratories,Hospital Laboratory,Laboratory, Hospital
D009426 Netherlands Country located in EUROPE. It is bordered by the NORTH SEA, BELGIUM, and GERMANY. Constituent areas are Aruba, Curacao, and Sint Maarten, formerly included in the NETHERLANDS ANTILLES. Holland,Kingdom of the Netherlands
D011786 Quality Control A system for verifying and maintaining a desired level of quality in a product or process by careful planning, use of proper equipment, continued inspection, and corrective action as required. (Random House Unabridged Dictionary, 2d ed) Control, Quality,Controls, Quality,Quality Controls
D012015 Reference Standards A basis of value established for the measure of quantity, weight, extent or quality, e.g. weight standards, standard solutions, methods, techniques, and procedures used in diagnosis and therapy. Standard Preparations,Standards, Reference,Preparations, Standard,Standardization,Standards,Preparation, Standard,Reference Standard,Standard Preparation,Standard, Reference
D003429 Cross Reactions Serological reactions in which an antiserum against one antigen reacts with a non-identical but closely related antigen. Cross Reaction,Reaction, Cross,Reactions, Cross
D005434 Flow Cytometry Technique using an instrument system for making, processing, and displaying one or more measurements on individual cells obtained from a cell suspension. Cells are usually stained with one or more fluorescent dyes specific to cell components of interest, e.g., DNA, and fluorescence of each cell is measured as it rapidly transverses the excitation beam (laser or mercury arc lamp). Fluorescence provides a quantitative measure of various biochemical and biophysical properties of the cell, as well as a basis for cell sorting. Other measurable optical parameters include light absorption and light scattering, the latter being applicable to the measurement of cell size, shape, density, granularity, and stain uptake. Cytofluorometry, Flow,Cytometry, Flow,Flow Microfluorimetry,Fluorescence-Activated Cell Sorting,Microfluorometry, Flow,Cell Sorting, Fluorescence-Activated,Cell Sortings, Fluorescence-Activated,Cytofluorometries, Flow,Cytometries, Flow,Flow Cytofluorometries,Flow Cytofluorometry,Flow Cytometries,Flow Microfluorometries,Flow Microfluorometry,Fluorescence Activated Cell Sorting,Fluorescence-Activated Cell Sortings,Microfluorimetry, Flow,Microfluorometries, Flow,Sorting, Fluorescence-Activated Cell,Sortings, Fluorescence-Activated Cell
D006650 Histocompatibility Testing Identification of the major histocompatibility antigens of transplant DONORS and potential recipients, usually by serological tests. Donor and recipient pairs should be of identical ABO blood group, and in addition should be matched as closely as possible for HISTOCOMPATIBILITY ANTIGENS in order to minimize the likelihood of allograft rejection. (King, Dictionary of Genetics, 4th ed) Crossmatching, Tissue,HLA Typing,Tissue Typing,Crossmatchings, Tissue,HLA Typings,Histocompatibility Testings,Testing, Histocompatibility,Testings, Histocompatibility,Tissue Crossmatching,Tissue Crossmatchings,Tissue Typings,Typing, HLA,Typing, Tissue,Typings, HLA,Typings, Tissue
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000911 Antibodies, Monoclonal Antibodies produced by a single clone of cells. Monoclonal Antibodies,Monoclonal Antibody,Antibody, Monoclonal
D001530 Belgium A country in Western Europe, bordering the North Sea, between France and the Netherlands. The capital is Brussels.

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