Anti-red Blood Cell Antibodies Research Articles

Isolation of human anti-red blood cell antibodies by repertoire cloning.

Isolation of human anti-red blood cell antibodies by repertoire cloning.

Kinetics of the autologous red cell agglutination test

The kinetics of the autologous red cell agglutination test for detecting circulating antibodies to HIV-1 were studied. Two monoclonal anti-red blood cell antibodies (1C3/86 and 10F7MN) were used to construct Fab-peptide conjugates for the test. Both antibodies recognize glycophorin α on the surface of erythrocytes by immunoprecipitation or immunoblotting techniques. The number of binding sites, association and dissociation constants of 1C3/86 Fab and 10F7MN Fab′ fragments were determined ( n = 4.80 × 10 5 sites/erythrocyte, K a = 0.43 × 10 7 M −1, K d = 23 × 10 −8 M for 1C3/86, n = 4.66 × 10 5 sites/erythrocyte, K a = 1.05 × 10 7 M −1, K d = 9.5 × 10 −8 M for 10F7MN. The binding studies were performed under the same conditions as the autologous red blood cell agglutination test. When 0.9 μg of anti-glycophorin Fab was added to 10 μl of blood 0.25 μg of 1C3/86 Fab was bound whereas 0.29 μg for 10F7MN Fab′ was bound. Antibody binding reached a plateau after 2 min and once bound did not exchange with unbound Fab over the time scale of the test. The binding of the anti-peptide antibody (cross-linking antibody) was also complete within 2 min. Addition of approximately 0.1 μg of anti-peptide antibody gave half a maximal agglutination score. This is equivalent to 10 μg/ml circulating antibody. Under the agglutination test conditions, Fab-peptide conjugate was bound to 14% of available glycophorin molecules. Half maximal agglutination occurred when approximately 1.1% of the bound Fab-peptide conjugates were cross-linked. A maximum agglutination score of four occurred in the presence of 1 μg of anti-peptide antibody equivalent to 100 μg/ml circulating antibody whereas an agglutination score of 1+ was elicited by only 0.032 μg anti-peptide antibody and involved the cross-linking of approximately 160 glycophorin per red cell.

Differential Usage of an Ig Heavy Chain Variable Region Gene by Human B-Cell Tumors

A monoclonal anti-idiotypic antibody has been raised that recognizes Igs with heavy chains encoded by a member of the VH4 family, the VH4-21 gene segment. The idiotope (Id) is detectable on a high percentage of early B cells in fetal spleen, and is expressed by certain autoantibodies, particularly cold-reactive anti-red blood cell antibodies. Therefore, it was of interest to investigate usage of this VH gene by neoplastic B cells; 81 chronic lymphocytic leukemias (CLLs) involving CD5+ B cells and 62 B-cell lymphomas of varying histologic type have been analyzed. The Id was expressed by only 3 of 81 (3.7%) of the CLLs, indicating a relatively low usage by these tumors. In contrast, the Id was expressed by 9 of 62 (14.5%) of the lymphomas across a range of histologic types, indicating a differential use of the VH4-21 gene among B-cell neoplasms. For three of the Id-positive lymphomas, each of a different histologic class, the nucleotide sequence of the tumor-derived VH gene was determined; the VH4-21 gene was identified, as expected. The sequence from the CLL was identical to the germline sequence, and the marginal zone lymphoma showed only 3 nucleotide changes, 2 of which gave rise to amino acid substitutions. In contrast, the sequence from the follicular lymphoma showed 29 nucleotide changes giving rise to 14 amino acid substitutions, which were scattered among the CDR and FW regions.

Enzyme linked immunosorbent assay for detection of anti-red blood cell(RBC) antibodies using RBC ghost protein.

Enzyme linked immunosorbent assay for detection of anti-red blood cell(RBC) antibodies using RBC ghost protein.

A modified radio-immune antiglobulin test for the screening of anti-red blood cell antibodies in hybridoma supernatants.

A radio-immune antiglobulin test was developed using mouse monoclonal anti-human A red blood cell (RBC) antibodies, and applied to the screening and study of hybridoma supernatants containing anti-human RBC antibodies. The latter antibodies were provided either as known monoclonal antibodies from culture supernatants and ascitic fluids, or as culture supernatants to be screened. The technical feasibility of the test was greatly facilitated by the use of remova-well disposable polystyrene plates, multichannel pipettes, and multiple-plate centrifuge holders. The sensitivity of the test and its relevance for the screening of hybridoma anti-human RBC antibodies is discussed.

ABO-incompatible marrow transplants.

We analyzed data from 81 ABO-incompatible marrow transplants performed between 1972 and 1980. Patients with high anti-red blood cell antibody levels pretransplant had a significantly increased probability of antibody return post-transplant. This was true for IgG and for IgM as independent variables. However, the return of antibody post-transplant had no effect on engraftment, survival, or blood product requirements posttransplant. Patients who received ABO-incompatible marrow had a similar time to engraftment, survival, and incidence of rejection and graft-versus-host disease as patients receiving ABo-compatible transplants. Neither pretransplant treatment regimen nor the development of graft-versus-host disease had an effect on post-transplant antibody levels. ABO-incompatible marrow transplants can be performed with no greater morbidity or mortality than ABO-compatible grafts. The procedures for antibody removal may be useful in other organ transplant settings.

Thyroid-Stimulating Antibodies in Patients with Autoimmune Disorders

A radioreceptor assay was used to measure thyroid-stimulating antibody (TSAb) in 1) patients with Graves' disease with untreated hyperthyroidism, selected for absence of clinically significant eye disease; 2) patients with Graves' ophthalmopathy, with and without previously treated hyperthyroidism; 3) patients with other thyroid disorders; 4) patients with other autoimmune disorders; and 5) normal subjects. TSAb was detected in 14 of 15 (93%) patients with Graves' hyperthyroidism and in 10 of 16 (63%) patients with Graves' ophthalmopathy. Of the patients with Graves' ophthalmopathy, TSAb was detected in 9 of 10 patients who had once been hyperthyroid and in only 1 of 6 patients who had never been hyperthyroid (euthyroid Graves' disease). TSAb was detected in 1 patient with idiopathic Addison's disease (autoimmune adrenalitis) and in 1 patient with juvenile diabetes mellitus (both of whom were euthyroid), and borderline levels were found in 1 patient with Sjögren's syndrome and 1 patient with methyldopa-induced antired blood cell antibodies. TSAb was not detected in normal subjects or patients with other thyroid disorders. The conclusions are: 1) the test is very useful in the diagnosis of Graves' disease; 2) Graves' eye disease may be a frequently associated but separate disorder; and 3) because TSAb may be present in some euthyroid patients with other autoimmune disorders, TSAb production may occur primarily because of a disorder in the immune system.

Detection of anti-red blood cell antibodies by treatment of the cells with neuraminidase.

Neuraminidase treated RBC were used to detect incomplete unbound anti RBC antibodies in the plasma of patients with immune diseases. 80% (48 out of 60) of the examined patients gave positive results with this method while all the 50 normal controls were negative. The indirect antiglobulin test was negative in all the patients. In the direct antiglobulin test only two RBC samples, whether treated with neuraminidase or not, gave positive results. The present study indicates that the assay of plasma on neuraminidase treated RBC may serve as a sensitive method for the detection of the presence of anti RBC antibodies in various immunological diseases.

Antibody response in patients with acute nonlymphocytic leukemia.

Lymphocytotoxic (LCT) and anti-red blood cell (ABO) antibodies were measured serially in adult patients with acute nonlymphocytic leukemia (ANLL) receiving induction chemotherapy. The antigenic stimulus was provided by multiple platelet transfusions, many of which were ABO incompatible. Comparison of pretherapy titers 4-6 weeks into therapy shows that 50% (19/38) of patients became LCT positive (cytotoxicity against greater than 10% of panel of cells) and 54% (19/35) had increases in ABO titers (greater than 2 tube dilution). A total of 66% of patients had significant rises in at least one antibody. ABO and LCT titers tended to vary in parallel although exceptions were noted. The development of anti) remission rate or duration, 3) type of therapy, 4) number of platelet transfusions, 5) time relationship between the antigenic stimulus and the initiation of cytotoxic therapy, and 6) skin test reactivity. Antibody responders tended to have higher pretreatment lymphocyte counts. The ability to develop a secondary antibody response does not appear to be a major prognostic factor in ANLL.

The Effects of Concentrated Eluted Anti-Red Blood Cell Antibodies on the in Vivo Survival of Normal Red Blood Cells

Abstract 1. A method has been described for the preparation and sterilization of a concentrated eluate from human red cell stroma. 2. Red cells sensitized by such an eluate prepared from normal control red cells showed entirely normal in vivo survival, as did cells sensitized by eluate from anti-H coated cells. 3. Sensitization of red cells by concentrated eluates from a patient with Coombs-negative acquired hemolytic anemia and from a patient with Coombs-positive acquired hemolytic anemia did not cause significant alteration in the in vivo survival of the red cells. 4. Red cells sensitized by the concentrated eluate from anti-D sensitized cells disappeared from the recipient’s circulation very rapidly and were sequestered in the spleen, indicating preservation of the physiologic properties of the antibody throughout the elution, concentration and sterilization procedures.