From incontinence to confidence.
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Abdomen↗
Biomedical subjects
S T Johnson
Publications and source records attributed to S T Johnson.
Prenatal genotyping of the Duffy blood group system by allele-specific polymerase chain reaction.
Maternal allo-immunization to antigens of the Duffy blood group system can result in haemolytic disease of the newborn (HDN), therefore, the application of allele-specific polymerase chain reaction (ASPCR) for prenatal genotyping of the Duffy antigen system to identify pregnancies at risk for HDN was evaluated. Oligonucleotide primers were designed for ASPCR of FYA, FYB and nullFY alleles. A validation study was performed using DNA isolated from 94 serotyped whole blood samples and 8 amniocentesis samples. A concordance rate of 100 per cent was observed between serotyping and ASPCR detection of the FYA, FYB and nullFY alleles. This assay is particularly useful for rapid genotyping of fetal amniotic cells to identify pregnancies at risk for HDN due to maternal fetal incompatibilities within the Duffy blood group system.
Alleles↗
Immunoglobulin isotype identification in red cell antibodies using flow cytometry.
BACKGROUND: Identifying the isotype of an immunoglobulin (IgM vs. IgG) detected in a patient sample is especially important in anticipating the risk of hemolytic disease of the newborn. Currently, 2-mercaptoethanol (2-ME) treatment of a sample is used in the authors' laboratory to degrade IgM, and this is followed by retesting. This method has multiple drawbacks. The purpose of this study was to develop a flow cytometry (FC) assay that would replace the 2-ME treatment protocol (2-ME treatment). STUDY DESIGN AND METHODS: A preliminary FC assay was developed, modified, and refined through the use of stock antibodies. Then, 10 samples containing antibodies were tested in parallel by the FC assay and 2-ME treatment. RESULTS: When a 10-unit mean channel fluorescence change was used as an index of a positive result, the FC assay detected all isotypes identified by 2-ME treatment. The FC assay was also able to identify mixtures of isotypes. One antibody that had not reacted in conventional agglutination testing was detected by the FC assay. The amount of fluorescence and the agglutinating strength of the antibody did not parallel each other. In one case, this discrepancy may have reflected an antibody that was primarily IgA. CONCLUSIONS: The FC assay appears to be as accurate as 2-ME treatment in differentiating IgG from IgM. The FC assay produces a positive endpoint for both isotypes, will identify IgA, requires less sample, and has no odor.
Antibodies, Anti-Idiotypic↗
Prenatal genotyping of Jk(a) and Jk(b) of the human Kidd blood group system by allele-specific polymerase chain reaction.
An allele-specific polymerase chain reaction (ASPCR) assay for prenatal genotyping of the Kidd antigen system in order to identify pregnancies at risk for haemolytic disease of the newborn (HDN) was developed. Oligonucleotide primers were designed for ASPCR of JKA and JKB. A validation study was performed using DNA isolated from 54 serotyped whole blood samples and 8 amniocentesis samples. A concordance rate of 100 per cent was observed between serotyping and ASPCR detection of the JKA and JKB alleles. Experiments were conducted to quantify the maternal contamination that could be tolerated in Kidd ASPCR assays. The sensitivity of this assay ranged from 0.2 per cent when detecting the presence of JKB and JKA background, to 2 per cent for detecting the presence of JKA in a JKB background. This sensitive assay is particularly useful for rapid genotyping of fetal amniotic cells to identify pregnancies at risk for HDN due to incompatibilities within the Kidd blood group system.
Adult↗
Survey on pretransfusion testing.
BACKGROUND: Hospitals and blood centers throughout the United States use a variety of reagents and methods to perform pretransfusion testing. A survey was developed to determine the reagents and methods in use and their relative prevalence in different work settings. STUDY DESIGN AND METHODS: A national survey on pretransfusion testing was conducted. Surveys were distributed to state and regional blood bank associations, which then distributed them to hospitals and blood centers within their region. In most instances, the blood centers distributed the survey to the local hospitals. Completed surveys were returned to the authors for review, and all information was entered into a database for analysis. RESULTS: Analysis of the data shows that the majority of blood banks use monoclonal reagents for ABO testing and monoclonal-polyclonal blended reagents for Rh testing. The data show that anti-IgG and polyclonal antihuman globulin reagents are used almost equally for antibody screening (detection) tests and that most blood banks use a three-cell antibody-screening test. Slightly more than 50 percent of hospitals use an immediate-spin crossmatch in the absence of unexpected antibodies. CONCLUSION: A number of approved reagents and methods are used by blood bank laboratories for pretransfusion testing. Facility size (number of beds) and type tend to influence the choice of methods and reagents employed. This survey provides an opportunity for blood bank laboratories to compare their current practices with those of their peers.
Blood Banks↗
Sensitivity to a metabolite of diclofenac as a cause of acute immune hemolytic anemia.
A 75-year-old woman taking the nonsteroidal anti-inflammatory drug diclofenac (DCF) presented with acute Coombs-positive hemolytic anemia and subsequently developed renal failure. A drug-dependent antibody specific for red blood cells (RBCs) could not be demonstrated by in vitro testing with DCF. However, her serum was found to contain an IgM antibody that reacted strongly with RBCs in the presence of urine from any of four subjects who had ingested DCF. The active substance in urine was isolated, subjected to high-performance liquid chromatographic (HPLC) analysis, and found to be a glucuronide conjugate of a known DCF metabolite, 4'-hydroxydiclofenac (4'-OH DCF). Negative results were obtained with four other DCF metabolites. Two 4'-OH DCF glucuronides were synthesized in vitro using a liver microsomal system. One promoted agglutination of normal RBCs by the patient's serum and was identified as the glucuronide ester of 4'-OH DCF by proton nuclear magnetic resonance (NMR) analysis. Studies with a panel of RBCs showed that the patient's antibody reacted preferentially with the e antigen of the Rh system. Acute immune hemolytic anemia in this patient appears to have been caused by sensitization to DCF modified by 4' hydroxylation and glucuronidation. This is the first reported example of immune cytopenia caused by sensitivity to a glucuronide conjugate of a drug metabolite. Since glucuronidation is a common pathway of drug metabolism, studies of the frequency with which glucuronide derivatives of primary medications cause immune cytopenia seem warranted.
Aged↗
Serum antibody responses of cattle following experimental infection with Escherichia coli O157:H7.
Oral inoculation of calves and steers with 10(10) CFU of Escherichia coli O157:H7 induced prompt and sustained increases in serum antibodies to O157 lipopolysaccharide. Neutralizing antibodies to verotoxin 1 also increased rapidly in most steers but more gradually in calves. None of the animals developed neutralizing antibodies to verotoxin 2. These serological responses were not correlated with elimination of infection in calves or steers or protection of calves against reinfection with the same strain.
Animals↗
Detection of Escherichia coli O157:H7 in meat by an enzyme-linked immunosorbent assay, EHEC-Tek.
Investigation of the specificity of an enzyme-linked immunosorbent assay (ELISA) for detection of Escherichia coli O157:H7 in raw meats (EHEC-Tek; Organon Teknika Corp.) revealed that the target antigens of the detection reagent, monoclonal antibody 4E8C12, were present in numerous serotypes of E. coli and that their ELISA reactivity was influenced by bile salts, acriflavine, and heat. The specificity of the ELISA was improved by a modified test protocol incorporating immunocapture.
Blotting, Western↗
Detecting fetomaternal hemorrhage: a comparison of five methods.
Appropriate postpartum administration of Rh immune globulin relies on sensitive detection and accurate quantitation of fetomaternal hemorrhage (FMH). Recently, the microscopic Du test (micro Du) enhanced with polyethylene glycol (PEG Du) and flow cytometry (FC) have been advocated for this purpose. Three qualitative methods (micro Du, rosette test, and PEG Du) and two quantitative methods (acid elution and FC) for assessing FMH were evaluated with particular attention given to PEG Du and FC. In vitro studies comprised 10 series of dilutions of D+ cord cells in D- adult cells to yield D+ cell concentrations of 0.06, 0.12, 0.25, 0.50, 0.75, 1.0, and 2.0 percent. Additionally, 26 postpartum samples were tested. Of the qualitative techniques, the micro Du test was the least sensitive with 20 percent false-negative results occurring at 0.5 percent fetal cells. The PEG Du test was only slightly more sensitive and offered no clinical advantage. The rosette test was the most sensitive, consistently detecting fetal cells at concentrations of 0.25 percent or greater. FC and acid elution showed similar results, with good correlation obtained between measured and expected quantities of fetal cells (r = 0.99 and 0.96, respectively). One of 26 postpartum samples was positive by all screening techniques; acid elution and FC detected 0.3-percent concentrations of fetal cells and 0.17-percent concentrations of D+ cells, respectively. Although acid elution is a more commonly used method for quantitating FMH, FC offers an acceptable alternative that is capable of analyzing large numbers of cells with objectivity and reproducibility.
Coombs Test↗
Cyanate: a potential uremic toxin.
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Biological Transport↗