Multicenter evaluation of the flow cytometry T-cell crossmatch: results from the American Society of Histocompatibility and Immunogenetics-College of American Pathologists proficiency testing program.

BACKGROUND: The performance characteristics and interlaboratory comparisons of the T-cell flow cytometry crossmatch remain largely unknown. METHODS: This study was performed using data from the ASHI-CAP proficiency testing program. Four unknown sera and two unknown cells were sent to participating laboratories twice a year for 4 years. RESULTS: In one survey in which different crossmatch techniques were compared, flow cytometry was slightly more sensitive than the antiglobulin method and considerably more sensitive than direct cytotoxicity. However, the proportion of participants in any given survey detecting antibodies in all sera expected to be positive was 50-60% and has not changed over the years. Failure to detect antibodies correlated with low antibody concentration, diluting the unknown serum by the testing laboratory, and with the instrument used. False positive results with normal sera were infrequent. Fluorescence intensity values were not standardized and were highly variable, but when fluorescence units reported by individual laboratories were divided by their own positive-negative cutoff values, results from different centers were more comparable. In general, fluorescence-to-cutoff ratios >5 correlated with complement binding activity, whereas values <5 denoted concentrations below those required to fix complement. CONCLUSIONS: Flow cytometry, as used by most centers, is highly sensitive and allows relative antibody quantitation. Furthermore, the data define objective parameters that may help to standardize the test and improve its predictive value in clinical transplantation.

A randomized trial comparing double-drug and triple-drug therapy in primary cadaveric renal transplants.

A controlled trial was carried out in 209 primary cadaveric renal transplants to compare the effects of cyclosporine and steroids (double therapy) with those of cyclosporine in lower initial dose, azathioprine, and steroids (triple therapy). Patients have been followed 1-36 months since transplantation. Actuarial two-year graft survival (double 74%, triple 76%) and two-year patient survival (double 90%, triple 93%) were similar for both groups. Further analysis of particular risk factors including age, diabetes, HLA matching, acute renal failure, and use of sequential Minnesota antilymphocyte globulin in patients with delayed graft function also showed similar outcomes with both immunosuppressive regimens. Initial hospitalization time, rate of rejection, incidence of serious infection, and rate of rehospitalization were not different. Mean CsA doses and mean trough whole-blood levels were higher in double-therapy patients at hospital discharge but not by three months posttransplant. There were no differences between the two groups in iothalamate clearance at any time. Hypertension was more frequent six months posttransplant in the triple-therapy group (p less than 0.05). Thus, similar results were obtained with both regimens, and except for hypertension no regimen appeared to have increased side effects up to three years posttransplant.

Effective cell support by repeat plateletpheresis of related donors on the IBM 2997 cell separator.

Forty-three non-HLA-matched donors provided 83 plateletpheresis products for 20 thrombocytopenic patients during a six-month period. The platelet product yield was 4.2 x 10(11) collected in 90 min. There was 26% cellular depletion of donor platelets and 20% depletion of donor lymphocyte per procedure. Males had a significantly greater lymphocyte depletion: 24% compared with 14% for females (P less than 0.05). There was no significant cellular depletion seen in 16 donors who underwent from two of a maximum of nine procedures. For these 16 donors, the time interval between procedures was a minimum of three days and a maximum of 100 days. Twelve refractory oncology patients received 49 plateletpheresis transfusions from 26 related donors. The mean corrected one-hour posttransfusion platelet increment was 18,300, and the mean corrected 20-hour posttransfusion platelet increment was 13,000. The results indicate that non-HLA-typed related plateletpheresis donors can safely undergo multiple procedures with the IBM 2997 Cell Separator and can effectively support their thrombocytopenic relatives who are unresponsive to random donor platelets.

Estimation of anti-D IgG in red blood cell eluates using the specific radioactivity of 125I-labeled IgG: effect of unlabeled, cytophilic IgG.

The specific radioactivity of conventionally prepared 125I IgG anti-D eluates is significantly less (from 1/5 to 1/20) than that of the 125I IgG fraction used to prepare the eluate. This discrepancy is due to the release of unlabeled, cytophilic IgG from normal red blood cells during eluate preparation and does not represent an underestimation of the eluate anti-D IgG content. Cytophilic IgG content of eluates plays an important role in reducing the nonimmunologic binding of labeled antibody IgG. The results justify the assumption used in numerous studies that the specific radioactivity of 125I IgG fractions can be used to provide a valid estimate of the anti-D IgG content of eluates.

Production of allo-anti-Ena by an individual whose red blood cells carry some Ena antigen.

We recently described an individual whose red blood cells appear to carry a hybrid MNSs sialoglycoprotein (SGP). The MN-derived portion of that SGP carries at least two determinants defined by some examples of antibodies that have been called anti-Ena. However, the red blood cells lack a different determinant that is defined by other examples of anti-Ena. This individual has now formed an anti-Ena antibody that reacts with the portion of Ena that her red blood cells lack, but not with the Ena determinants that have been shown to be carried on MN SGP. It is not yet clear whether the portion of Ena that her red blood cells lack and that her antibody defines is MN SGP-borne. The findings in this case provide further support for our conclusions that the terms "Ena" and "anti-Ena," as previously used, describe heterogeneous groups of antigens and antibodies.

Another individual (J.R.) whose red blood cells appear to carry a hybrid MNSs sialoglycoprotein.

The serum of J.R. contains anti-Wrb and her red blood cells are of the phenotype Wr(a-b-). Evidence was obtained that suggested that her cells totally lack normal MN and Ss sialoglycoproteins (SGPs), and carry instead an abnormal SGP, which is likely to be a hybrid SGP resembling the MN SGP in its outer portion and the Ss SGP in its inner portion. Although the apparent hybrid SGPs of J.R. and the MiV homozygote are virtually indistinguishable in terms of their electrophoretic mobility (apparent molecular weight 40,000) and staining characteristics, they are not identical. That of J.R. is associated with a weak M and increased S antigen, while that of the MiV homozygote is associated with a very weak N, a greatly exalted s, and the rare antigen, Hil. Like the study on the blood of the MiV homozygote, serologic studies on the red blood cells of J.R. have revealed considerable heterogeneity of what have previously been called the Ena antigen and anti-Ena antibodies.