Four examples of anti-TSEN and three of TSEN-positive erythrocytes.

BACKGROUND: TSEN (MNS33) is a low-incidence antigen in the MNS blood group system encoded by hybrid glycophorin genes. TSEN is expressed by a unique amino acid sequence that results from the junction of GPA(58) to GPB(27), if the GPB carries S antigen. Until this study, only one example of anti-TSEN had been found. Antibody screening red blood cells (RBCs) positive for both S and s (ref. No. C873) reacted with four patient sera. Initially, the RBCs had been typed as S+s+, but later were typed as S-s+ in another laboratory. Two other RBC samples, one from a volunteer blood donor (D.L.), the other from a patient whose serum contained anti-En(a)FR (J.S.), also gave anomalous results when tested with anti-S. We suspected the presence of TSEN-positive hybrids on all three RBC samples. MATERIALS AND METHODS: Reactive sera (O.B., E.C., S.K., R.F.) were tested against RBCs with normal MNS phenotypes and with TSEN-positive RBCs. The RBCs of D.L., J.S. and C873 were tested with anti-S whose reactivity with S+s+ TSEN+ RBCs had been established previously, and with the original example of anti-TSEN. Immunoblotting was performed on the C873, D.L. and J.S. RBC membranes using a monoclonal antibody to an epitope common to both glycophorin A and glycophorin B. RESULTS: The sera from O.B., E.C., S.K. and R.F. were strongly reactive on the indirect antiglobulin test with TSEN+ RBCs. The RBCs of C873, D.L. and J.S. were typed as TSEN+. Immunoblotting pattern of D.L. and C873 were consistent with TSEN heterozygotes, while that of J.S. was consistent with a TSEN homozygote. CONCLUSIONS: Based on the estimated number of screening events with C873 RBCs, the incidence of anti-TSEN is approximately 1 in 20,000 sera. The antibody is found in patients with and without documented exposure to allogeneic RBCs. All known examples of anti-TSEN are IgG, but their clinical significance is not known.

Capture-S, a nontreponemal solid-phase erythrocyte adherence assay for serological detection of syphilis.

A solid-phase erythrocyte adherence assay has been developed for the serological detection of reagin antibodies in syphilis. Capture-S (Immucor, Inc., Norcross, Ga.) is a nontreponemal, qualitative screening test for the detection of immunoglobulin G (IgG) and IgM antilipid antibodies in serum or plasma samples from blood donors. The Capture-S assay utilizes a modified Venereal Disease Research Laboratory antigen bound to microtitration wells and anti-IgG- plus anti-IgM-coated indicator erythrocytes as the detection system. The Capture-S assay was evaluated at six separate sites on 10,942 specimens. For patient samples of clinically diagnosed syphilis categories (n = 366), the Capture-S assay yielded a sensitivity of 80.7% versus 80.3% for the rapid plasma reagin (RPR) card test (Becton Dickinson Microbiology Systems, Cockeysville, Md.). In comparative experiments on patient and donor samples (n = 10,222), the Capture-S assay demonstrated a sensitivity of 94% compared to 91.2% for the RPR card test. The Capture-S and RPR card tests produced essentially equivalent specificities of 99.2% and 99.3%, respectively, for this sample population. For five test sites, the Capture-S and RPR card test demonstrated a 98.3% agreement (10,085 of 10,264) of test results. These evaluations indicate that the Capture-S compares favorably to the RPR card test in assay sensitivity and specificity, with the added benefits of ease of use, accommodation of high-volume testing, and potential for automation.

Detection of tube agglutination 37 degrees C-only antibodies by solid-phase red cell adherence.

There are no published data on the detection of tube agglutination (TA) 37 degrees C-only antibodies by solid-phase (SP) red cell adherence assays using anti-IgG-coated indicator red cells. Thirteen examples of TA 37 degrees C-only antibodies were tested by conventional SP methods. Four TA 37 degrees C-only antibodies failed to react by SP. Three were anti- Lea, considered clinically insignificant, and one was anti-E, an antibody of potential clinical significance. The remaining nine TA 37 degrees C-only antibodies reacted by SP, including three anti-c, two anti- D, two anti-E, one anti-N, and one anti-M. The anti-M reacted with indicator red cells that lacked the red cell antigen and failed to react with IgG-coated indicator red cells whose anti-IgG component had been neutralized, indicating the antibody contained an IgG component. Two anti-D and one anti-c continued to react in an SP test using neutralized anti-IgG antigen-positive indicator red cells, i.e., indicator binding independent of antiglobulin, suggesting an IgM nature to these antibodies. Therefore, many TA 37 degrees C-only antibodies can be detected by SP either through detection of an IgG component by the anti-IgG of the indicator red cells, or through IgM crosslinking of antigen-positive indicator red cells to antigen-positive SP reagent red cell membranes.

Detection of Lewis, P1, and some MNS blood group system antibodies by a solid phase assay.

Some solid phase red cell adherence (SPRCA) assays are designed to detect IgG antibodies to red blood cell (RBC) antigens. These assays use anti-IgG-coated red cells as the indicator. It is reported that most antibodies to Lea, Leb, P1, M, and N fail to react by solid phase (SP), presumably because they are IgM antibodies. Those detected are assumed to be IgG. In one year, during routine testing using SPRCA to screen patients for intended RBC transfusion, 28 of 59 such examples were found to react: anti-Lea(9), -Leb(1), -M(14), -N(1), and -P1(3). A study was undertaken to determine if reactivity was due to crosslinking by IgM antibodies of antigen-positive indicator RBCs to antigen-positive reagent RBC monolayers, or due to detection of IgG antibodies. Antibodies were tested according to standard SP protocols, except where IgG-neutralized indicator RBCs were substituted for anti-IgG-active indicator cells. The 59 samples were retested with antigen-positive and antigen-negative indicator RBCs. Only 5 of 59 reacted optimally when antigen-positive indicator cells were used: anti-Lea(2), -Leb(1), -M(1), and -N(1). The reactions of all antibodies were abolished when the anti-IgG component of the indicator was neutralized by soluble IgG. These findings show that detection of most Lewis, P1, M, and N antibodies by SPRCA is dependent on the presence of an IgG antibody in the serum.

Antibody detection errors due to acidic or unbuffered saline.

Isotonic saline solutions, buffered with potassium phosphate or sodium phosphate salts, were evaluated in parallel with unbuffered saline to determine if they improved antibody detection by solid phase red cell adherence or hemagglutination methods. Saline buffered to a pH of 7.0 to 7.5, when used to suspend red cells or to wash sensitized red cells in preparation for the antiglobulin test, produced the best positive solid phase and hemagglutination results. The pH range of commercially prepared blood bank saline (unbuffered) was found to be 5.8 to 6.8, far lower than the desired pH for optimum antibody detection. In the case of solid phase assays employing intact, immobilized reagent red cells, saline with a pH of 7.0 to 7.5 also eliminated falsely positive results due to the dissociation of red cell monolayers from the solid support surface that occurred in the presence of unbuffered or acidic saline. These findings indicate that unbuffered isotonic saline should not be used in solid phase- or hemagglutination-based antibody detection tests. It is recommended that phosphate-buffered saline at a pH of 7.0 to 7.5 be employed.

High-titer, low-avidity (HTLA) antibodies and antigens: a review.

Antibodies that react to HTLA characteristics cause difficulties in serologic testing because of the weak reactions they produce in the indirect antiglobulin test. Those specificities that are more frequently encountered (anti-Yka, -McCa, -Kna, -Ch) are directed toward antigens of high incidence in both the white and black populations. They have not been shown to cause significant destruction of transfused antigen-positive red cells. The antibodies create problems in serologic tests because the reactions they produce interfere with the identification of reactions due to other, clinically significant antibodies.

Recovery of costs for special procedures done in reference laboratories.

In recent years, blood bank laboratories have been encouraged to look at special procedures that might generate needed revenues. Laboratories that elect to offer special procedures to recover costs or to generate revenue should evaluate each test carefully Areas to be considered are the projected demand for the test, proficiency of present laboratory staff, cost of reagents and equipment, and assay time relative to labor cost.

Studies on the blood of an MsHe/MSu proposita and her family. Serological evidence that Henshaw-producing genes do not code for the 'N' antigen.

The serum of a black woman has been found to contain a potent alloanti-N that reacted in direct agglutination tests with the 'N' antigen carried on Ss-active sialoglycoprotein. Thus far, such antibodies have been observed only in the sera of MSu/MSu individuals. However, our proposita had red cells that lacked 'N'. Her blood type was M+, N-, S-, s+, U+, He+. Results of family studies indicate that she is of the genotype MsHe/MSu. Our findings are consistent with recently reported data on the structure of the Henshaw antigen, which is located on a Ss-active sialoglycoprotein that does not carry 'N'.

Difficulty in LW typing as revealed by a family study.

A family is described in which two members of the second generation are of the phenotype LW3. In the course of the investigation the mother of the LW3 propositus was at first believed to be phenotypically LW3 as well. Eventually, it was shown that she is, in fact, phenotypically LW2 but that her R-1 (no D), LWlw, genotype had resulted in less LW being present on her red blood cells than is expected in LW2 persons. This reduced level of LW could not be detected with one example of anti-LW made by an LW3 individual.

Autoantibodies mimicking alloantibodies.

A patient with myelofibrosis, who has produced many red blood cell autoantibodies, is described. Although the patient is phenotypically R1R1 (CDe/CDe), eluates made from his red blood cells have consistently contained what appeared to be anti-E, and more recently another antibody that appeared to be anti-c. In in vitro experiments we have shown that the "anti-E" and "anti-c" can be totally adsorbed by E-negative and c-negative red blood cells, respectively. We conclude that the two antibodies have quite different specificities from those indicated by simple antibody identification studies, and that both are more closely related to the anti-Hr series of antibodies than to anti-E or anti-c.

Failure to demonstrate dosage of U antigen.

Tests in which 11 examples of anti-U were used in titration studies against the red blood cells of 9 obligate Uu heterozygotes, from 4 unrelated families, and random Negro and Caucasian donors (many of whom were of the presumptive UU genotype) have failed to demonstrate any dosage of the U antigen.

A new example of anti-LW and further studies on heterogeneity of the system.

A case is reported in which an LW3 woman formed anti-LW. It is shown that the case represents the "genetic" and not the "transient" type of red blood cell LW antigen depletion. Studies have confirmed the quantitative difference of LW antigen on LW3 and LW4 red blood cells and the fact that Rhnull individuals produce the only totally LW-negative red blood cells. The difficulties encountered in the identification of anti-LW, because of the LW3 and LW4 states, are described. It is clear, from this study, that if random donors are typed for LW using anti-LW made by an LW4 indvidual, those who are LW3 will be classed as unremarkable LW-positives.