[Irregular blood group antibodies during pregnancy: screening is mandatory]. / Irregulaire-bloedgroepantistoffen tijdens de zwangerschap: meten is weten.

During pregnancy irregular blood group antibodies, originating either from earlier pregnancies or from blood transfusions, may severely jeopardize both mother and child. Three patients are described with pregnancy-associated blood group incompatibility. In one case of Kell antagonism a previous child had reportedly died of cot death, but in retrospect it had most probably suffered from erythroblastosis fetalis as a result of anti Kell antibodies. In the second case, a twin pregnancy, the diagnosis of neonatal haemolytic anaemia on the basis of blood group incompatibility with a very rare antibody (anti-Kpb) had been established in the previous child. No precautions had been taken during this pregnancy, putting both mother and children at risk. All three children recovered, the twins after repeated transfusion of Kpb-free erythrocytes. The described cases emphasize the importance of being informed about the presence of antibodies during pregnancy. Such information can only be obtained by assessing the antibody status during pregnancy. In the Netherlands, the screening of all pregnant women for the presence of irregular antibodies was introduced last year.

Erythrocyte aging in the demented elderly: a fluctuating process?

Measurement of erythrocyte aging parameters in patients with dementia indicates that an Alzheimer-related disturbance of the erythrocyte aging process may not be detectable until in the more advanced stages of the disease. Also, a strong fluctuation in the values of erythrocyte aging parameters, over a period of 15 months, was observed in patients with dementia, but not in age-matched control donors. This fluctuation was independent of the type and stage of dementia, and its cause remains to be elucidated. Such variability hampers the use of erythrocyte aging characteristics for the diagnosis of dementia. On the other hand, the aging-related erythrocyte IgG content may be a sensitive biomarker for disturbed systemic homeostasis in the elderly.

Erythrocyte aging characteristics in elderly individuals with beginning dementia.

An increase in erythrocyte-bound IgG and enhanced breakdown of the erythrocyte anion exchanger band 3, characteristics of normal erythrocyte aging are observed in old, healthy individuals when compared with young donors. These findings indicate that the rate of cellular aging increases with organismal aging. Results from previous studies on the same parameters have suggested that the erythrocyte aging process is disturbed in patients in advanced stages of Alzheimer type dementia, and in individuals with Down's syndrome who show no signs of dementia. In this study we find no changes in erythrocyte aging parameters in old individuals in beginning stages of dementia of various etiologies. We conclude that, in general, characteristics of disturbed erythrocyte aging cannot serve as presymptomatic markers of Alzheimer-type dementia.

Flow cytometric method for the routine follow-up of red cell populations after bone marrow transplantation.

A simple and sensitive flow cytometric method is presented for the quantitation of erythrocyte subpopulations. Cells were indirectly labelled using human antisera (mostly crude) and fluorescein isothiocyanate (FITC)-conjugated anti-human IgG Fab. The method was evaluated by analysing artificial mixtures of blood group antigens A, B, D, E, c, K and Fy(a). Intra-assay coefficients of variation were found to be 7.2% for 1% D-positive mixtures and 2.2% for 10% D-positive mixtures: the inter-assay coefficients of variation were 9.5% and 6.0% respectively. The sensitivity of the method was found to be 0.31%. The method has shown to be suitable for the routine follow-up of patients after allogeneic bone marrow transplantation (BMT).

Blood group chimerism in human multiple births is not rare.

Twin blood group chimerism seems to be very rare in humans. The 30-40 previously reported cases usually were found by mere coincidence during routine blood grouping in hospitals or blood banks. Usually in these cases frank blood group mixtures of, for example, 50/50%, 25/75%, or 5/95% at most were seen. Smaller percentages are very difficult to notice during routine work-up. Using a sensitive fluorescence technique (sensitivity > 0.01%) we detected blood group chimerism in 32/415 (8%) twin pairs and 12/57 (21%) triplet pairs, respectively, which is a higher incidence than reported previously.

Erythrocyte repopulation after major ABO incompatible transplantation with lymphocyte-depleted bone marrow.

Forty-four out of 258 allogeneic BMT were performed across the major ABO barrier. Donor erythrocyte repopulation could be evaluated in 30 cases. Fifty-eight patients transplanted with an ABO compatible or minor incompatible graft served as the control group. All patients received a marrow graft depleted of lymphocytes by counterflow centrifugation. Less than 10(8) residual erythrocytes were present in the graft. Cyclosporin A was used as immunoprophylaxis after transplantation. Erythrocyte repopulation was measured using a fluorescent microsphere method. An adapted transfusion policy was applied. Eight out of 30 patients (27%) with major ABO incompatibility had no detectable donor erythrocytes 2 months after BMT. Up to 3 months after BMT donor erythrocyte repopulation was significantly delayed in the ABO incompatible group (P < or = 0.03). Significantly more erythrocyte transfusions were required in the ABO incompatible group (P < 0.001). Six patients with blood group O (20%) developed pure red cell aplasia which resolved in five without therapeutic intervention. In these six patients anti-A antibody titers were persistently high the first 3 months after BMT. This was in contrast with 22 patients with timely recovery of erythropoiesis in whom anti-A and anti-B antibody titers showed a steady decrease after BMT. The incidence of immunohematological complications in these patients who received a lymphocyte depleted major ABO incompatible graft is similar (20%) to the incidence reported in the literature. Serious morbidity related to major ABO incompatibility did not occur.

Influence of aging and neurodegenerative disease on changes in band 3-like proteins in white blood cells.

Fluorescent microspheres were used to measure antibody-induced capping of leukocyte membrane proteins that are immunologically related to band 3, the anion exchanger of the erythrocyte. The degree of capping was found to increase with donor age. Surface labeling and capping characteristics of cells from healthy, age-matched controls were not different from those from patients with Alzheimer's disease, multi-infarct dementia, and Down's syndrome. Immunoblots, however, indicated increased expression and/or breakdown of band 3-like proteins in leukocytes from patients when compared with young and old control donors. These findings emphasize the possible involvement of band 3-like proteins of nucleated cells in aging and disease.

Erythrocyte membrane changes of individuals with Down's syndrome in various stages of Alzheimer-type dementia.

An increase in erythrocyte-bound IgG, enhanced breakdown of band 3, and changes in erythrocyte anion transport characteristics are observed in individuals with Down's Syndrome. We interpret these data as indicative for a disturbance of normal erythrocyte aging. A comparison of three groups of individuals with Down's Syndrome in various stages of dementia of the Alzheimer type did not reveal a correlation between the erythrocyte membrane changes and age or stage of dementia. Considering previously obtained data on patients with senile dementia of the Alzheimer type (7), these results may signify that parameters of disturbed erythrocyte aging represent presymptomatic markers of Alzheimer-type dementia.

Influence of the conditioning regimen on erythrocyte chimerism, graft-versus-host disease and relapse after allogeneic transplantation with lymphocyte depleted marrow.

Three different conditioning regimens were applied to 144 patients undergoing allogeneic bone marrow transplantation (BMT) with HLA identical sibling marrow, depleted of lymphocytes by counterflow centrifugation. All regimens consisted of cyclophosphamide and fractionated total body irradiation (TBI). In 49 patients treated with regimen A the total TBI dose was 9 Gy. In regimen B the dose rate of TBI was increased and anthracyclines were added (n = 65). Thirty patients received regimen C with a total TBI dose of 12 Gy but no anthracyclines. The different conditioning regimens did not influence the percentage of patients with detectable recipient CFU-GM prior to infusion of donor marrow. The incidences of mixed erythrocyte chimerism at 6 months after BMT were 73, 33 and 20% for regimens A, B and C respectively. The conditioning regimen influenced significantly mixed erythrocyte chimerism from 6 to 24 months after BMT. Both age and the conditioning regimen influenced significantly the incidence of acute graft-versus-host disease (GVHD) (p = 0.017 and 0.0001 respectively). Acute GVHD greater than or equal to I occurred in 15, 29 and 77% of the patients treated with regimens A, B and C respectively. The incidence of acute and chronic GVHD was significantly higher in complete donor chimeras than in mixed chimeras (p less than 0.001 and p less than 0.01). The probability of relapse was 43% in 32 and 18% in 43 good risk patients treated with regimens A and B respectively (p = 0.07). Longer follow-up is needed to draw conclusions about relapse in regimen C.(ABSTRACT TRUNCATED AT 250 WORDS)

Erythrocyte membrane characteristics indicate abnormal cellular aging in patients with Alzheimer's disease.

Erythrocytes from patients with Alzheimer's disease show signs of disturbance of the normal cellular aging process. Immunoblotting of erythrocyte membrane proteins from Alzheimer patients reveals increased breakdown of the anion transport protein band 3 in a majority of the cells, similar to what is observed in only a very small cell population during normal aging. These structural changes are accompanied by changes in anion transport characteristics, but the latter partially deviate from those observed during normal aging. The amount of erythrocyte-bound immunoglobulin G, the most direct and relevant parameter of erythrocyte aging, is significantly increased in Alzheimer patients relative to age-matched, healthy donors and to patients with multi-infarct dementia. These data indicate accelerated molecular breakdown of band 3 and premature appearance of senescent cell characteristics in erythrocytes from Alzheimer patients, and support the hypothesis that abnormal cellular aging may be involved in the etiology of the Alzheimer-specific pathology.

Evaluation of the polyethylene glycol antiglobulin test for detection of red blood cell antibodies.

The polyethylene glycol (PEG) antiglobulin test (AGT) was evaluated by testing antibodies in the sera of 362 different patients and by screening of 4,685 random patient sera with both the PEG AGT and the albumin AGT. Agglutination in the PEG method was stronger in 254 (70.2%), identical in 99 (27.3%) and weaker in 9 (2.5%) of the 363 sera. Mean end-point titers of the antibodies measured with the PEG method (n = 137) were 2-3 dilutions higher than with the albumin method. In sera with anti-K and anti-Fyb antibodies the mean titer in the PEG method was only one dilution higher. Antibodies within the rhesus system were also tested with a two-stage enzyme test. Agglutination in the PEG method was stronger in 93 (50.3%), identical in 64 (34.6%) and weaker in 28 (15.1%) of the 185 sera. Mean antibody titers (n = 76) in the PEG method were 1-3 dilution steps higher for the various specificities than in the enzyme test. In 92 (2.0%) of the 4,685 sera screened in the PEG method, specific antibodies were detected; with the albumin method in 74 (1.6%) sera. The antibodies only detected with the PEG method had the specificities anti-D,-C,-Cw,-c,E,-K,-Jka and -M. Not detected by the PEG method were anti-Lea antibodies in one of the sera. It is concluded that the PEG AGT is a sensitive technique for the detection and identification of red cell antibodies and that it is superior to the albumin AGT.

Host and donor erythrocyte repopulation patterns after allogeneic bone marrow transplantation analysed with antibody-coated fluorescent microspheres.

Analysis of erythrocyte populations, using red blood cell antigen differences between host and donor as marker, was performed with a sensitive fluorescent microsphere assay to monitor marrow engraftment and mixed red cell chimaerism after allogeneic bone marrow transplantation (BMT). An adapted transfusion policy, using marker negative erythrocyte transfusions, was required for this analysis. In all patients the marrow graft was depleted of lymphocytes by counterflow centrifugation. Thirty-seven patients were evaluable for donor repopulation at one or more points in the first 6 months after BMT. At 0.5 month donor erythrocytes were detectable in 19 out of 22 patients. At 6 months donor erythrocytes were detectable in 100% of the evaluable patients. In the first 3 months after BMT the average donor erythrocyte repopulation in recipients of major ABO mismatched transplantations was delayed. Thirty-eight patients were evaluable for chimaerism at 6 months or later after BMT. A high incidence of mixed red cell chimaerism was observed varying from 50% to 71% at different points of analysis. Mixed red cell chimaerism with low percentages (less than 1%) of host cells was not related with relapse, nor did high percentages (greater than 10% of host cells necessarily indicate relapse.

A fluorescent microsphere method for the investigation of erythrocyte chimaerism after allogeneic bone marrow transplantation using antigenic differences.

A method for the investigation of erythrocyte chimaerism in patients after bone marrow transplantation (BMT) has been developed using fluorescent microspheres coated with anti-human IgG. Blood group antigens different in patient and donor were used as marker. The specificity and reliability of this method was evaluated measuring artificial mixtures of blood group positive and negative red cells. Red cell antigens tested were D, c, K, Fya, Fyb, Jka, Jkb, S and s. Mean linear regression lines for mixtures with percentages positive cells ranging from 0.01 to 1% and 1 to 100% were 0.91X-0.03 (r = 0.99) and 1.00X-0.05 (r = 0.99), respectively. The sensitivity level of the assay was one positive cell per 10,000 blood group negative cells. In negative control samples (n = 15) a mean percentage positive cells was found of 0.002 +/- 0.004 (SD). Intra- and inter-assay coefficients of variation were 4.9 and 5.3% for mixtures of 10%, and were 11.1 and 24.6% for mixtures of 0.1% blood group positive cells. It is concluded that the described method can be used both to establish the take of donor marrow in an early phase after transplantation and to investigate mixed erythrocyte chimaerism long after BMT.

Aldosterone secretion rate: radioimmunoassay versus double isotope dilution derivative assay.

A simple and reliable radioimmunoassay for the measurement of the aldosterone secretion rate has been developed. In this method the urinary pH 1-hydrolyzed aldosterone is acetylated to its 21-monoacetate and purified by a single paper chromatogram. The isolated aldosterone 21-monoacetate is radioimmunoassayed using the N.I.H. antiserum lot 088. From the value obtained and the tritium amount present in the eluate the specific activity is calculated. The following results disclosed the reliability of the method: 1. The values obtained were in perfect agreement with values obtained by a double isotope assay. 2. The aldosterone secretion rate values calculated did not change upon extension of chromatographic steps. 3. Immunochromatographic analysis showed a sharp peak in the aldosterone 21-monoacetate area without significant background immunoreactivity. 4. The steroid concentration measured decreased linearly with dilution of the paper eluate. 5. Identical values were obtained with three antisera, raised against different aldosterone immunogens. In our laboratory no reliable results could be obtained when the conversion to the aldosterone 21-monoacetate was omitted and the specific activity was calculated after radioimmunoassay of chromatographically purified, pH 1-hydrolyzed, aldosterone per se. In that case the values obtained were systematically lower than the double isotope values, a problem which is discussed.