Biomedical subjects

R F Brunhouse

Publications and source records attributed to R F Brunhouse.

4 recordsLinked to original sources

PubMed1997

A whole blood alternative to traditional methods for CD4+ T lymphocyte determination.

Flow cytometry, the method of choice for determining subset cell populations, requires sophisticated instrumentation and highly trained operators. An alternative method for subpopulation determinations is presented that is fast, simple, and readily interpreted for absolute subset counts. This method uses antibody labeled microspheres added to whole blood and after lysing, read on a hemocytometer using an ordinary light microscope. The total time of testing is less than 10 min with results comparable to that obtained using a flow cytometer and a hematology analyzer to calculate absolute subpopulation values. This method is demonstrated for CD4+ lymphocytes which have considerable value in following HIV-infected individuals.

Adult↗

PubMed1995

Lymphocyte subset determination using a hematology analyzer.

Anti-CD4 antibody (T4)-coated microspheres were used to label CD4 cells in whole blood. The mixture was lysed and analyzed by a modified Coulter VCS hematology analyzer, which differentiated microsphere-labeled cells by a change in Coulter volume, conductance, and light scatter. %CD3+/CD4+ fluorescent values from a profile were compared to %CD4 values using the VCS-microsphere method. CD3 gating was used to exclude CD4+ monocytes from the 90LS-FALS lymphocyte gate. The results correlated well (R = 0.996). The percentage of CD4+ lymphocytes from profile scatterplots and VCS scatterplots showed a line of regression close to the equivalence line (n = 76, slope = 0.96) when CD3 gating was used for the profile. These results suggest that CD3 gating, though necessary for 90LS-FALS scatterplots, may not be necessary for volume-conductance-light scatterplots.

Antibody Specificity↗

PubMed1978

Guinea pig immunoglobulin light chain isotypes. I. Separation of kappa and lambda chains and the identification of three isotypes of the lambda chain constant homology region.

The separation of intact kappa chain and two fragments comprising lambda (lambda) chain from immunoglobulin light chain pools isolated from strain 13 guinea pigs is achieved by cyanogen bromide digestion and gel filtration before and after reductive clevage of disulfide bonds. The smaller lambda chain fragment derives from the original carboxyl-terminus of lambda chain. The partial sequence of component tryptic and thermolytic peptides of this thirty-nine residue fragment allowed its complete sequence to be deduced. Three isotypic forms of this lambda chain constant region fragment, distinguished by four residue positions showing alternative amino acids, are found expressed by guinea pigs. Each of these three isotypes is more homologous to the other two than to lambda chains of any other species. The distribution of amino acid substitutions differentiating these isotypes further supports the view that lambda chain isotypes arose in guinea pigs, and in several other species, independently by gene duplication.

Amino Acids↗

PubMed1978

Guinea pig immunoglobulin light chain isotypes. II. The preferential expression of lambda chain-bearing anti-phenyltrimethylammonium antibodies is associated with restricted variable region expression, but not with any alteration in the proportion of lambda chain constant region isotypes, relative to normal immunoglobulins.

The inbred strain 13 guinea pig expresses at least three isotypes of immunoglobulin (Ig) lambda chain. The proportion of these isotypes in normal Ig can be estimated by isolating a constant (C) region cyanogen bromide peptide from lambda chains and quantitatively analyzing its component tryptic peptides. Anti-phenyltrimethylammonium antibody, when analyzed in this way, was found to contain the same proportion of each lambda chain C region isotype as did normal Ig. The specific antibody light (L) chain pools were found to contain predominantly lambda chains, although these are the minority type in guinea pig normal Ig L chain pools. Moreover, amino-terminal, 5-pyrrolidone-2-carboxylic acid-containing peptides isolated from antibody lambda chains show no demonstrable sequence heterogeneity, while their homologs isolated from normal lambda chains are markedly heterogeneous. That a restriction in heterogeneity of variable region framework residue positions is not associated with any alteration in the proportion of C region isotypes in these antibody lambda chains, implies that translocation of all guinea pig lambda chain variable region genes to the isotypic C region genes occurs in a random way.

Animals↗