Flow Immunomagenetic Beads For Detecting The BCR-ABL Fusion Proteins Of Chronic Myelogenous Leukemia Study On The Methodology

ObjectivesTo explore streaming immunomagnetic beads in the detection bydifferent methods of control through the use of different methods ofcytogenetics, reverse transcriptase polymerase chain reaction, flowimmunomagnetic beads method to detect the BCR-ABL fusion gene inchronic myeloid leukemia, the gene evaluation methodology.Methods1.Detected by cytogenetics, reverse transcriptase polymerase chainreaction, flow immunomagnetic beads of the same group of K562celllines(1) cytogeneticsRapid culture of k562, by preventing mitosis, hypotonic treatment,pre-fixed, fixed, centrifugation, drops piece step technique usingthermal denaturation Giemsa G-banding karyotype analysis of metaphase cells analyzed.(2) reverse transcriptase polymerase chain reactionExtraction the RNA of K562cell line and cDNA synthesis, theproducts wereamplification, further analysis of the amplified product.(3) Flow immunomagenetic beadsExtract of K562cells cultured pretreatment steps of the fusion proteinBCR-ABL in K562cell strains were detected, dissolved of proteinmicrospheres mark on the machine.2.o evaluation the specificity of the flow immunomagnetic beadsdetection of BCR-ABL fusion proteinCulturing HEL cell lines，though the three different methods，suchas cytogenetics, PCR and flow immunomagnetic beads to detect and toevaluate its specificity.3.To evaluation the sensitivity of the flow immunomagnetic beadsdetection of BCR-ABL fusion proteinAfter the culture of K562cells then diluted at differentconcentrations (100%,10%,1%,0.5%,0.2%,0.05%,0%), detectedthe diluted K562cells,in order to observe CBA sensitivity in the detectionof BCR-ABL fusion protein.Results1、The mean fluorescence intensity of BCR-ABL fusion protein in marrow cell from BCR-ABL negative protein was83.84±49.8.2、The sensitivity detection of BCR-ABL fusion protein detectiondilution test results by flow immunomagnetic beads show that in normalhuman peripheral blood mononuclear cells were diluted to1%in theK562cell line, continued to show positive results.3、Flow immunomagnetic beads assay specificity of the BCR-ABLfusion protein detection normal human peripheral blood mononuclearcells、HEL cell lines and K562cells were detected.Just the K562celllines display a strong PE fluorescent signal, rather than the expression ofthe BCR-ABL fusion protein in peripheral blood mononuclear cells andHEL cell lines show negative PE fluorescence signal.4、Select the30specimens of K562cells were detected, results showthat it has better MFI,8specimens out the30specimens， after labeledbeads in darkroom show that the MFI decreased after20hours, but stilldetected by the positive results.ConclusionsDetecte the sensitivity and specificity of the BCR-ABL fusion proteinin K562cell line by Flow immunomagnetic beads，it’s show that theresults are good, with its accurate, simple, rapid establishment of themethodology for future clinical promote the CBA to the foundation.