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PubMed2003

[Development of an anti-rTNF-alpha chimeric antibody].

AIM: To develop an human-mouse chimeric antibody against rTNF-alpha. METHODS: Expression vector of human-mouse chimeric antibody CZ12 gene was constructed by using VL and VH genes of mAb Z12 with neutralizing activity, and then COS7 cells were transfected. Supernatant containing CZ12 was detected by RT-PCR, ELISA, Western blot and in-vitro neutralization assay, respectively. RESULTS: The RT-PCR analysis showed that there was the transcription of human-mouse chimeric antibody mRNA in the transfected COS7 cells. ELISA and Western blot detection showed that the CZ12 could recognize specifically TNF-alpha. The in-vitro neutralization assay proved that CZ12 could neutralize the cytoxicity of TNF-alpha to L929 cells. CONCLUSION: human-mouse chimeric antibody CZ12 with neutralizing activity was successfully expressed in eukaryotic cells.

Animals↗
PubMed2004

[Construction of eukaryotic vector and expression of chimeric antibody against rh-bFGF].

AIM: To express a human-mouse chimeric antibody against rh-bFGF antigen in eukaryotic cells. METHODS: The VL and VH genes were amplified and cloned from the hybridoma 1F11 secreting anti-rh-bFGF mouse monoclonal antibody (mAb) and the CL and CH genes were amplified and cloned from the plasmid pMDHC and pMDLC. They were inserted into eukaryotic expression vectors after sequencing. The recombinant plasmids were then transformed into CHO-dhfr- cells for expression. The humanization and antigen specificity of expressed products were identified by indirect ELISA. The relative molecular mass (Mr) of expressed products was determined by SDS-PAGE. RESULTS: The cloned antibody genes were identified to be functional by sequencing. The chimeric antibody could be detected in the culture supernatant of transformed CHO cells. The humanization and specificity of the chimeric antibody to rh-bFGF were confirmed by ELISA. CONCLUSION: The mouse-human chimeric antibody against rh-bFGF has been expressed successfully in eukaryotic cells, which lays the foundation for its further clinical research.

Amino Acid Sequence↗
PubMed1992

Differential effects of a murine and chimeric mouse/human anti-interleukin-2 receptor antibody on human T-cell proliferation.

The preference for interleukin-2 receptor (IL-2R) expression on activated, compared with resting T lymphocytes makes the IL-2R a promising target for selective immunosuppressive therapy. To increase the potential therapeutic effectiveness of anti-IL-2R monoclonals, a chimeric mouse/human variant was constructed from Ig genes isolated from a murine anti-human IL-2R hybridoma cell line, designated AHT54. AHT54 binds to the same or spatially related epitope as IL-2 on the p55 protein that constitutes the low- and high-affinity forms of IL-2R. Although the murine and chimeric AHT54 antibodies inhibited cell-surface binding of IL-2 to the same extent, the chimeric antibodies containing a human IgG1 constant region had substantially more anti-proliferative activity than their murine IgG1 counterparts. Our results indicated that the human constant region of the chimeric antibodies interacted more efficiently than the murine constant region with effector components present in peripheral blood mononuclear cells (PBMC).

Animals↗
PubMed2004

Short tandem repeat technology has diverse applications: individual identification, phylogenetic reconstruction and chimerism based post haematopoietic stem cell transplantation graft monitoring.

BACKGROUND: Short Tandem Repeat (STR) loci are widely considered to be effective for variety of applications including forensic applications, phylogenetic reconstruction and chimerism based post Haematopoietic Stem Cell Transplantation (HSCT) graft monitoring. For each application, specific sets of STR loci are used. AIMS: In the present study, we have attempted to use same set of STR loci for varied purposes based on their efficacy and informativity. SETTINGS AND DESIGN: Population and patient based study. MATERIALS AND METHODS: We have analyzed 5 STR loci--vWA, Tho1, FES, F13 and TPOX in 1000 North Indians. All five markers were also analyzed for chimerism based graft monitoring after HSCT in 42 HLA matched pair of patient-donor to predict the outcome of transplantation. STATISTICAL ANALYSIS: The analysis was done for Hardy Weinberg equilibrium (HWE), Heterozygosity, Polymorphism information content (PIC) and Power of Exclusion and Phylogenetic assessment. RESULTS AND CONCLUSIONS: High allelic variability in term of Heterozygosity (0.68-0.76), PIC (0.66-0.74) and high Power of exclusion (0.28-0.38) indicating high forensic utility. The ensuing PC plots finely resolved three basal clusters corresponding to three geo-ethnic groups of African, Orientals, and Caucasians. In post HSCT chimerism analysis, it was found that together these markers were informative in 38 pairs (98%) and were able to predict the chimerism status successfully. There is a possibility that these STR loci along with forensic and phylogenetic importance, can predict the outcome of HSCT successfully.

Gene Frequency↗
PubMed2004

[Potential of cdc25A-Fas chimeric expression vector in inducing apoptosis of Tca8113 cells in vitro].

BACKGROUND & OBJECTIVE: Previous studies have revealed the close relationship between Fas/FasL pathway and carcinogenesis of oral squamous cell carcinoma (OSCC). This study was designed to explore the potential of cdc25A-Fas chimeric expression vector in inducing apoptosis of human OSCC cell line Tca8113. METHODS: The 2 chimeric expression vector pAdTrack-CMV-cdc25A-Fas (pCCF), and pAdTrack-cdc25A-Fas (pCF) were constructed by gene engineering, pCCF, pCF, and the control plasmid pAdTrack-CMV were transfected into Tca8113 cells by liposome, respectively. The transfection efficiency was presented by the expression of the report gene, green fluorescence protein (GFP). The mRNA and protein levels of Fas were determined by Northern blot analysis, reverse transcription-polymerase chain reaction (RT-PCR), Western blot analysis, and immunohistochemistry methods. The apoptosis of transfected Tca8113 cells was analyzed by techniques of DNA agarose gel electrophoresis, TUNEL, Annexin V label, and flow cytometry (FCM). RESULTS: (1) The chimeric expression vectors pCCF, and pCF were successfully transfected into Tca8113 cells and the maximum transfection (15%) was observed at the 5th-7th day after transfection. (2) Up-regulation of Fas expression was detected at the 3rd day after transfection in pCCF, and pCF transfection groups. At 3rd, 5th, and 7th day after transfection, Fas protein was found to express on membrane and in plasma of transfected Tca8113 cells with distinct morphology of partial apoptosis. (3) From 2.5 days after transfection, early apoptosis had been observed in pCCF, and pCF transfection groups. At 3rd day, the increased apoptosis index (AI,=25%) of pCCF, and pCF transfection groups was significant higher than that of control group (P< 0.05), whereas there was no significant difference in AI between pCCF transfection group and pCF transfection group (P >0.05). (4) FCM analysis showed that the peak of GFP-expression cells was identical to that of the apoptotic cells. CONCLUSION: The cdc25A-Fas chimeric expression vector was able to initiate the apoptosis of Tca8113 cells by up-regulating Fas expression. This result indicated that the 27 bp cdc25A fragment can be designed as a cis-regulatory element to modulate the effects of C-Myc/Max in cellular proliferation and apoptosis.

Apoptosis↗
PubMed2004

[Application of sequential and quantitative analysis of donor chimerism in donor lymphocyte infusion].

In order to study the value of sequential and quantitative analysis of chimerism in determination of optional time of donor lymphocyte infusion (DLI) and prediction of efficacy of DLI, six patients with leukemias who relapsed or failed of engraftment were treated with DLI. Serial and quantitative analyses of donor chimerism (DC) both prior to and following DLI were performed by multiplex PCR amplification of STR markers (STR-PCR) and capillary electrophoresis with fluorescence detection. The results showed that at the time of relapse or graft rejection, STR-PCR indicated the decreasing donor chimerism in all six patients, at levels ranging from 27.3% to 85.7%. The declining value of DC (<90%) was detected in four patients at 26 days before relapse or graft rejection diagnosed clinically. Therefore the decrease of value of DC can be identified the high risk of relapse or graft failure and can be used to guide DLI implementation at early stage. In this study the clinical response were seen in two patients, the value of DC in these patients increased with convertion to a predominant donor profile (>90%) or converted to stable FDC shortly after DLI, while in the patients without clinical response, the level of DC decreased persistently or declined after transient increase. Three patients without response received second DLI. It is concluded that the monitoring of chimerism is proved to be a valuable to determine the optional time point of DLI and to early evaluate the efficacy of DLI. Furthermore, it can present a rational basis for treatment of intensification in the patients who did not respond to first-line DLI treatment.

Adolescent↗
PubMed1992

The uptake kinetics of chimeric oligodeoxynucleotide analogues in human leukaemia MOLT-4 cells.

Chimeric oligodeoxynucleotides with terminal nonionic methylphosphonate analogue sections and internal phosphodiester regions offer several advantages as antisense effectors over either structure alone. These include enhanced biological stability relative to all-phosphodiester molecules, increased activity in directing ribonuclease H mediated destruction of target RNA, increased specificity and reduced non-specific toxicity. However, another important parameter, the ability of these molecules to enter intact mammalian cells, has not previously been investigated. Therefore, oligodeoxynucleotides were tagged at their 5'-termini with fluorescein reporter groups and a detailed study of uptake kinetics in human leukaemia MOLT-4 cells undertaken by calibrated flow cytometry. Baseline measurements with all-phosphodiester and all-methylphosphonate molecules confirmed that uptake of oligodeoxynucleotides by intact cells is a highly inefficient process. The kinetic data were in agreement with previous reports of mechanisms of cell uptake involving receptor mediated endocytosis in the case of phosphodiester molecules and simple diffusion for methylphosphonates. Chimeric oligodeoxynucleotides exhibited saturable cell uptake kinetics similar to all-phosphodiester oligodeoxynucleotides, suggesting that uptake was receptor-mediated and distinct from concentration-dependent uptake of all-methylphosphonate molecules. Similarly, chimeric molecules were apparently confined to the endosomal compartment within cells. These results imply that reversible masking of the negative charges of the phosphodiester sections of chimeric oligodeoxynucleotides may be required to change the uptake mechanism back to simple diffusion and allow intracellular delivery to the site of the target RNA.

Amino Acid Sequence↗
PubMed2002

Employing chimeric xylanases to identify regions of an alkaline xylanase participating in enzyme activity at basic pH.

The xylanase A (XynA) from the alkaliphilic Bacillus halodurans C-125 and the xylanase B (XynB) from Clostridium stercorarium F9 were subdivided into four fragments at highly homologous regions present in their primary structures: an amino-terminal region (A or a), a region containing the putative proton donor (P or p), a region containing the putative catalytic nucleophile (N or n), and a carboxyl-terminal region (C or c). Six chimeric xylanases were constructed by the selective substitution of the four fragments using an overlapping PCR technique. Two of the six xylanases, APnc and Apnc (regions originating from XynA are denoted by upper case letters and those from XynB are denoted by lower case letters), were produced in Escherichia coli while the other four xylanases were obtained only as inclusion bodies. The APnc and Apnc chimeric enzymes were purified by column chromatography using Ni-NTA agarose and DEAE-Toyopearl. The respective pH and temperature stabilities of the purified enzymes were observed from pH 5.6 to 11.6 and up to 45 degrees C for APnc, and from pH 5.6 to 11.2 and up to 45 degrees C for Apnc. Thus, these enzymes were slightly less stable than the parental xylanases. An assessment of the pH-activity relationships for the chimeric xylanases employed p-nitrophenyl-beta-D-xylobioside as the substrate in determinations of the k(cat) values. The pK(a1) values for the APnc and Apnc chimeric enzymes were 4.3 and 4.2, respectively, which were almost identical to those for the parental xylanases. In contrast, the pK(a2) values obtained for APnc and Apnc were 9.1 and 8.5, respectively; these values fall between those for the parental xylanases, XynA (9.4) and XynB (7.8). These results indicate that the main regions necessary to maintain the high pK(a2) value of XynA locate in the A and P sections.

Journal Article↗
PubMed1991

Chimeric gamma-delta signal joints. Implications for the mechanism and regulation of T cell receptor gene rearrangement.

Rearrangement of Ag receptor genes requires recognition by the lymphocyte recombinase of heptamer-nonamer signal sequences followed by two endonucleolytic cleavages and two DNA ligations to form the coding and signal joints. The phenomenon of trans-rearrangement, in which Ag receptor gene segments located on different chromosomes recombine to yield chimeric products, provides an in vivo system in which to investigate the ability of the recombinase to carry out each of these functions in trans. Trans-rearrangements between TCRG and TCRD loci, similar in structure and frequency to those observed previously in human lymphoid tissues, were demonstrated in normal mouse thymus by PCR with crossed V gamma/J delta and V delta/J gamma primer pairs. A simple mechanistic model for trans-rearrangement was then tested. This model posits an ability of the recombinase to catalyze the formation of both coding and signal joints in trans and therefore predicts that trans-rearrangements will generate chimeric signal joints. In adult thymus, chimeric D delta 2-J gamma 1 and D delta 2-J gamma 2 signal joints, containing fused heptamer-nonamer sequences, could be detected by PCR and were each present at frequencies sufficient to account for a large proportion of the corresponding TCRG/TCRD trans-rearrangements. In agreement with the predictions of the model, chimeric signal joints were found as both linear chromosomal and circular episomal DNA. The results provide a framework for understanding the formation of chromosomal translocations in normal and neoplastic lymphoid cells and support the possibility of a looping mechanism for standard gene rearrangement. To test the form of regulation of TCRG rearrangement, the frequencies of specific signal joints from standard and trans-rearrangements were compared. Although J gamma 1 and J gamma 2 segments participated with equal frequency in trans-rearrangement with D delta 2, only the J gamma 1 segment participated in standard rearrangement with V gamma 5. The results suggest that V-J recombination in the TCRG locus is regulated directly at the DNA level by cis-acting constraints which do not affect the accessibility of individual TCRG gene segments to recombination in trans.

Animals↗
PubMed2006

Highly sensitive patient-specific real-time PCR SNP assay for chimerism monitoring after allogeneic stem cell transplantation.

Chimerism analysis after allogeneic stem cell transplantation (allo-SCT) is an important diagnostic tool for the documentation of engraftment, early detection of graft failure, and recurrence of the disease. Current assays rely on the genetic polymorphism between the donor and the recipient, and allow semiquantitative or quantitative analysis of chimerism. The most common method in use is based on the amplification of the short tandem repeats (STR). This method, with 1% to 5 sensitivity, is useful for the documentation of engraftment, but is insufficient for the detection of minimal residual disease or early relapse, when medical intervention is urgently needed. Recently, single-nucleotide polymorphism (SNP) has been suggested as an alternative, more accurate system to monitor chimerism. The purpose of our study was to develop an easy, economical, and sensitive method for the detection of chimerism following allo-SCT using the SNP technology. Our approach is based on SNP patient-specific quantitative real-time polymerase chain reaction (PCR) using nonlabeled primers. Our results show that this allele-specific SNP real-time PCR approach is sensitive, relatively cheap, and offers a fast and reliable assay for the monitoring of hematopoietic engraftment and for the detection of minimal residual disease in patients after allo-SCT.

Female↗
PubMed2006

[Construction and expression of chimeric anti-human CD20 monoclonal antibody].

AIM: To construct the eukaryotic expression vector of chimeric anti-human CD20 monoclonal antibody (mAb) and realize its expression. METHODS: The light- and heavy-chain genes were amplified from hybridoma cell line 1-28 secreting anti-human CD20 mAb by RT-PCR and were cloned to T vector and sequenced. Proteins of mAb 1-28 were separated by reducing SDS-PAGE. Light- and heavy-chain bands were excised from preparative gel, digested by trypsin, and subjected to peptide mass fingerprinting. Software Biolynx and pepeseq were used to evaluate the score of probability. Correctness of the light- and heavy-chain DNA sequences was verified by their protein sequences. Genes of V(H) and V(L) were amplified from T vector and cloned into chimeric antibody expression vector (pCMV-V(H) and pCMV-V(L)), generating the expression vectors of chimeric anti-human CD20 mAb (C1-28) including light chain expression vector C1-28L and heavy chain expression vector C1-28H. The two plasmids were co-transfected into 293T cells with Lipofectamine 2000. RT-PCR was used to detect the transcription at mRNA level. C1-28 expression was detected by Sandwich ELISA and Western blot methods. RESULTS: mAb 1-28's genes were successfully cloned and verified by peptide mass fingerprinting. Eukaryotic expression vectors of anti-human CD20 mAb were constructed and expressed in 293T cells with the expression amount reaching 257 mg/L and the molecular weight consistent with that of human IgG. CONCLUSION: These experiments lay solid foundation for further study on the role of chimeric CD20 antibody in the treatment of non-Hodgkin's lymphoma.

Antibodies, Monoclonal↗
PubMed2006

Effects of chimerism on graft-versus-host disease, disease recurrence, and survival after HLA-identical marrow transplantation in Iran.

BACKGROUND: The coexistence of recipient's and donor's hematopoietic systems after allogeneic marrow transplantation is called mixed chimerism. OBJECTIVE: The objective of this study was to evaluate the effects of MC on graft-versus-host disease (GVHD), disease recurrence, and survival after HLA identical marrow transplantation in a transplant center in Iran. METHODS: The association of MC with acute GVHD, disease recurrence, survival, and relapse-free survival was investigated in 91 patients who underwent either bone (n = 12) or peripheral blood (n = 79) HLA-identical marrow transplantation. Chimerism was assessed using multiplex amplification of short tandem repeats (STR). Patients had thalassemia (n = 19), acute myelogenous leukemia (AML) (n = 29), acute lymphocytic leukemia (ALL) (n = 20), chronic myelogenous leukemia (CML) (n = 18), and other diseases (n = 5). The median age was 21 (range: 3 - 50) years. There were 38 (42%) female and 53 (58%) male participants. Conditioning was made through busulfan plus cyclophosphamide in 34 patients; busulfan plus fludarabin in 51 patients; and busulfan plus fludarabin plus antithymocyte globulin in 6 patients. The median follow-up was 13 months. RESULTS: On day +30, complete chimerism (CC) was observed in 72 (79%) patients, MC in 15 (17%), and no chimerism in 4 patients. The incidence of acute GVHD was significantly (P = 0.01) lower in mixed chimeras than in complete chimeras. There was no significant difference in acute GVHD grade (I, II vs. III, IV) between the two groups. The incidence of relapse was 18%. There was no difference in relapse rate between MC and CC groups. Overall survival was 89%. There was no significant difference in the overall survival between MC and CC group (96% vs. 85%, respectively). Relapse-free survival was 80% that was not significantly different between the two groups. CONCLUSION: Despite some previous reports, we found no significant difference in the survival and relapse rates between MC and CC groups.

Adolescent↗
PubMed1991

Chimeric pneumococcal cell wall lytic enzymes reveal important physiological and evolutionary traits.

Two novel chimeric pneumococcal cell wall lytic enzymes, named LC7 and CL7, have been constructed by in vitro recombination of the lytA gene encoding the major autolysin (LYTA amidase) of Streptococcus pneumoniae, a choline-dependent enzyme, and the cpl7 gene encoding the CPL7 lysozyme of phage Cp-7, a choline-independent enzyme. In remarkable contrast with previous chimeric constructions, we fused here two genes that lack nucleotide homology. The CL7 enzyme, which contains the N-terminal domain of CPL7 and C-terminal domain of LYTA, exhibited a choline-dependent lysozyme activity. This experimental rearrangement of domains might mimic the process that have generated the choline-dependent CPL1 lysozyme of phage Cp-1 during evolution, providing additional support to the modular theory of protein evolution. The LC7 enzyme, built up by fusion of the N-terminal domain of LYTA and the C-terminal domain of CPL7, exhibited an amidase activity capable of degrading ethanolamine-containing cell walls. The chimeric amidase behaved as an autolytic enzyme when it was cloned and expressed in S. pneumoniae. The chimeric enzymes provided new insights on the mechanisms involved in regulation of the host pneumococcal autolysins and on the participation of these enzymes in the process of cell separation. Furthermore, our experimental approach confirmed the basic role of the C-terminal domains in substrate recognition and revealed the influence of these domains on the optimal pH for catalytic activity.

Base Sequence↗
PubMed1991

Amplification by the polymerase chain reaction of hypervariable regions of the human genome for evaluation of chimerism after bone marrow transplantation.

We combined the polymerase chain reaction (PCR) with oligonucleotide hybridization as a novel and sensitive technique to evaluate posttransplant chimerism. Specific oligonucleotides for hybridization were synthesized homologous to tandemly repetitive core sequences of regions with a variable number of tandem repeats (VNTRs). Polymorphisms at such loci result from allelic differences in the number of repeats. Primers flanking the repeat region of each of the corresponding VNTRs were used for amplification. Recipient and donor pretransplant DNA and recipient posttransplant DNA were amplified. The resultant fragments were analyzed after gel electrophoresis either by hybridization in-gel or after Southern transfer. To confirm our findings, we also performed standard assays of restriction fragment length polymorphisms (RFLPs). Evaluation of 13 selected cases indicated mixed chimerism (4), complete chimerism (5), recurrence of leukemia (2), and endogenous repopulation of hematopoiesis (2) after marrow transplantation. Sensitivity of the method was determined by mixing various proportions of recipient and donor DNA; the limit of detection of the minor component in a mixture was 0.1%. PCR data correlated with RFLP data in all cases except two in which PCR proved more sensitive than RFLP. PCR amplification of VNTRs combined with oligonucleotide hybridization is a novel technique for documenting posttransplant chimerism and has advantages over RFLP analysis: high sensitivity, use of small amounts of DNA (250 ng), ease of preparation of DNA, elimination of need for restriction enzymes, and the ability to complete studies in 2 days.

Base Sequence↗
PubMed2006

Generation of cloned and chimeric embryos/offspring using the new methods of animal biotechnology.

The article summarizes results of studies concerning: 1/ qualitative evaluation of pig nuclear donor cells to somatic cell cloning, 2/ developmental potency of sheep somatic cells to create chimera, 3/ efficient production of chicken chimera. The quality of nuclear donor cells is one of the most important factors to determine the efficiency of somatic cell cloning. Morphological criteria commonly used for qualitative evaluation of somatic cells may be insufficient for practical application in the cloning. Therefore, different types of somatic cells being the source of genomic DNA in the cloning procedure were analyzed on apoptosis with the use of live-DNA or plasma membrane fluorescent markers. It has been found that morphological criteria are a sufficient selection factor for qualitative evaluation of nuclear donor cells to somatic cell cloning. Developmental potencies of sheep somatic cells in embryos and chimeric animals were studied using blastocyst complementation test. Fetal fibroblasts stained with vital fluorescent dye and microsurgically placed in morulae or blastocysts were later identified in embryos cultured in vitro. Transfer of Polish merino blastocysts harbouring Heatherhead fibroblasts to recipient ewes brought about normal births at term. Newly-born animals were of merino appearance with dark patches on their noses, near the mouth and on their clovens. This overt chimerism shows that fetal fibroblasts introduced to sheep morulae/blastocysts revealed full developmental plasticity. To achieve the efficient production of chicken chimeras, the blastodermal cells from embryos of the donor breeds, (Green-legged Partridgelike breed or GPxAraucana) were transferred into the embryos of the recipient breed (White Leghorn), and the effect of chimerism on the selected reproductive and physiological traits of recipients was examined. Using the model which allowed identification of the chimerism at many loci, it has been found that 93.9% of the examined birds were chimeras. The effect of donor cells on the reproduction and physiology of the recipients was evident.

Animals↗
PubMed1991

Transfectomas expressing both secreted and membrane-bound forms of chimeric IgE with anti-viral specificity.

Because of the lack of a cell line expressing on surface and secreting human IgE of known Ag specificity, the construction of a transfectoma line possessing such properties would be useful for studying the roles of surface IgE and the effects of anti-IgE antibodies on IgE-producing B cells. Toward this goal, the human genomic DNA segment encompassing the two exons encoding the membrane anchor peptide of epsilon-chain and their flanking regions was sequenced. Hybrid epsilon and kappa genomic DNA comprising the C regions of human epsilon- and kappa-chains and the H and L chain V regions of the murine mAb BAT123, which reacts with the gp120 envelope protein of HIV-1, were constructed. Mammalian expression vectors containing these fusion genes were used to transfect murine myeloma Sp2/0 cells, and transfectants stably expressing on surface and secreting into culture medium chimeric IgE were obtained. The chimeric IgE showed identical Ag-binding properties as the murine mAb BAT123. Acting in concert with the specific peptide Ag polyvalently coupled to a protein carrier, the chimeric antibody could induce histamine release from human blood basophils. These results demonstrate the potential utility of the transfectoma cells and the chimeric IgE in studying the roles of membrane-bound IgE and effects of anti-IgE antibodies on IgE-producing B cells.

Amino Acid Sequence↗
PubMed2006

Flow cytometric evaluation of red blood cell chimerism after bone marrow transplantation in Iranian patients: a preliminary study.

The aim of this study was to evaluate mixed red cells population and red blood cell chimerism after hematopoietic stem cell transplantation. Red blood cell chimerism after hematopoietic stem cell transplantation was analyzed using a series of fluorescein isothiocyanate-conjugated monoclonal antibodies (BioAtlantic, France) directed against ABH, Rh (D, C, E, c, e), Kell, Duffy, Kidd, and Ss antigens on blood samples of 14 patients with hematologic disorders undergoing hematopoietic stem cell transplantation, by flow cytometric method on days 15, 30, and 60 after transplantation. All patients showed expression of donor red cell antigens within days 15 - 30 after hematopoietic stem cell transplantation. Graft versus host disease and ABO incompatibility did not affect the expression of chimerism. Flow cytometric analysis is a simple, accurate, and valuable test which is of significant help in monitoring chimerism in allogeneic hematopoietic stem cell transplantation.

ABO Blood-Group System↗
PubMed1991

[Application of Y-chromosome specific DNA analysis for detection of mixed hematopoietic chimerism after allogeneic bone marrow transplantation].

We report a case involving mixed hematopoietic chimerism after an allogeneic bone marrow transplantation (BMT) from a sex mismatched donor. A 31 year-old-man who was diagnosed as having chronic myelogenous leukemia in the accelerated phase received an allogenic BMT from his HLA-identical sister in March, 1989. To determine the mixed chimerism we used the Y-chromosome specific repeated sequence of DNA using a specific probe (PHY 10). The donor's DNA 3.5 kb band appeared in 1-10% of male DNA by Southern blot hybridization in the peripheral blood 21 days after BMT. The Y-chromosome DNA band decreased day by day, and disappeared 110 days after BMT. The Y-chromosome DNA band could be detected, even though few metaphases were obtained immediately after BMT. Thus this method is very sensitive for determining which cells contain the Y-chromosome, and is therefore useful for detecting mixed chimerism after sex-mismatched BMT. Using this method the clinical significance of mixed chimerism can be assessed.

Adult↗