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PubMed1981

Measurement of nanogram concentrations of IgG by mixed hemadsorption.

UNLABELLED: We have modified the mixed hemadsorption technique so as to minimize the non-specific adsorption of indicator cells. This is accomplished by: (1) introducing fetal calf serum at several crucial steps in the procedure, and (2) standardizing the selection of high-power fields to be counted so as to avoid erratic adherence patterns which occur around the well rim. Using these adjustments, IgG concentrations in the 12.5--400 ng/ml range can be measured accurately. SAMPLE SIZE: 2 microliter. Test time: 4 h. No radioisotope is required.

Animals↗
PubMed1990

Hemadsorption and fusion inhibition activities of hemagglutinin analyzed by vaccinia virus mutants.

Vaccinia virus IHD-J strain induces hemagglutinin (HA) on the surface membrane of infected cells and does not elicit cell-cell fusion (F-). We isolated 21 independent hemadsorption-negative (HAD-) mutant viruses from IHD-J and five HAD+ revertants from one of these mutants. Of the 21 mutants, 19 that synthesized either no or little HA at the cell surface caused cell-cell fusion (F+), whereas none of the five revertants that synthesized HA at the cell surface induced cell-cell fusion. Furthermore, anti-HA monoclonal antibody B2D10 induced extensive polykaryocytosis of IHD-J-infected cells and suppressed the ability of the IHD-J-infected cell extract to inhibit the polykaryocytosis induced by IHD-W. The other 2 of the 21 HAD- mutants, B1 and A2, which induced HAs at the cell surface, showed F- and F+ phenotype, respectively. The HA molecule of mutant B1 had a single amino acid substitution of Lys for Glu-121 in its extracellular domain, whereas that of mutant A2 had a single substitution mutation of Tyr for Cys-103. We conclude that the vaccinia HA is a fusion inhibition protein, that the active sites for the two activities reside separately in its extracellular domain, and that cysteine-103 is important in forming the proper tertiary structure of the protein to exert both activities.

Animals↗
PubMed1987

Solid phase reverse passive hemadsorption test for hepatitis B surface antigen.

A solid phase reverse passive hemadsorption test (SP-RPHAd) for hepatitis B surface antigen detection is described. It was compared with a commercial reverse passive hemagglutination assay (Hepatest, Wellcome, U.K.). SP-RPHAd is four-fold less expensive than Hepatest and undiluted sera can be used instead of eight-fold diluted sera without risk of non-specific hemagglutination. Thus, the threshold of detection is lowered to about 5 ng/ml by SP-RPHAd.

Enzyme-Linked Immunosorbent Assay↗
PubMed1990

Analysis of bovine parainfluenza-3 virus replication in bovine embryonic lung cells by indirect fluorescent antibody and hemadsorption assays.

The temporal manifestation of bovine parainfluenza-3 virus (BPI3V) proteins in the cytoplasm and on the surface of bovine embryonic lung (BEL) cells was characterized by indirect fluorescent antibody (IFA) and hemadsorption (HAd) assays. Intracellular proteins appeared earliest at 0.5 h post inoculation (p.i.) and the infection spread to virtually all cells by 48 h p.i. Viral proteins on the surface of cells were seen first at 16 h p.i., and by 48 h p.i. the entire cell monolayer was IFA positive. Hundred-fold less virus in the inoculum delayed appearance of the intracellular as well as cell-surface viral proteins by 24 h and allowed the infection of only about 1/3 of the cells by 48 h p.i. Kinetics of the development in the proportion of HAd-positive cells correlated with those of the surface fluorescence-positive cells. Morphology of the manifestation of BPI3V proteins is characterized by microphotography.

Animals↗
PubMedDate not supplied

Comparison of hemagglutination and hemadsorption tests for influenza detection.

Hemagglutination (HA) testing of the supernatant of primary rhesus monkey kidney cell culture tubes inoculated with clinical specimens was compared with hemadsorption (HAd) of the same tube for identifying the presence of hemagglutinin activity due to influenza. A total of 476 respiratory specimens were screened for influenza by this method. The 127 influenza or parainfluenza positive cultures showed 100% concordance between the two tests. All negative cultures were both also negative by HA and HAd. In contrast to the HAd method, HA testing was performed with less tube manipulation, less hands-on time, and less potential for cell culture tube contamination.

Animals↗
PubMed2002

A mixed hemadsorption assay for detection of cell surface binding anti-tumor antibodies in human sera.

A practical mixed hemadsorption assay (MHA) was developed for the detection of cell surface binding anti-tumor antigen antibodies (anti-TA Ab) in the human sera. The assay was compared to enzyme-linked immunosorbent assay (ELISA) and dot ELISA in the detection of the antibodies developed against GA733-2 antigen. SW1116 cell line and recombinant GA733-2E protein were used as targets in the MHA and solid phase immunoassays (SPIA), respectively. Sera were obtained from healthy donors and vaccinated-colon carcinoma patients. The sensitivity level of the MHA was very high for detection of anti-TA Ab in sera.

Antibodies, Neoplasm↗
PubMed1983

A hemadsorption method for detection of colonies of Haemophilus influenzae type b expressing fimbriae.

Although fimbriated variants of Haemophilus influenzae type b have recently been described, cultures of most clinical isolates contain only a small proportion of fimbriated forms. Because colonies of fimbriated and nonfimbriated cells are visually indistinguishable, a rapid, simple method was developed for the identification and quantitation of colonies of fimbriated H influenzae. This procedure, also applicable to other bacteria (for example, Escherichia coli), involves transferring the colonies from agar to nitrocellulose disks and incubating the disks in a suspension of red blood cells. Colonies that contain predominantly fimbriated bacteria bind the red blood cells and appear as red dots on the nitrocellulose. This nitrocellulose hemadsorption method is described, as well as its applicability for determining the proportion of fimbriated cells in a culture, the kinetics of enrichment of fimbriated forms during enrichment procedures, and the transition rate from the nonfimbriated to the fimbriated state.

Bacteriological Techniques↗
PubMed1977

Electron microscopic study of hemadsorption on vaccinia virus infected cells.

Hemadsorption (HAD) induced in HEp-2 cells infected with vaccinia virus was observed. In ultrathin sections, binding of 36 red blood cells (RBCs) was examined in detail and 3 types of HAD were observed: (1) direct and close binding of RBCs to infected HEp-2 cells (cyto-HAD) was observed in cross sections of 27 RBCs (2) binding of RBCs through microvilli of infected cells was found in 11 RBCs, and (3) five RBCs were distorted to form tentacle-like projections by which they were bound to the HEp-2 cell surface. Scanning electron microscopy revealed that more than 30% of the RBCs were bound to microvilli of vaccinia virus-infected HEp-2 cells, and that the number of microvilli twined round each RBC was over ten, RBCs were attached to certain microvilli through swollen sucker-like tips which were not observable in non-infected HEp-2 cells. RBCs sometimes revealed a polygonal shape at regions of binding to microvilli. Virion-mediated RBC HEp-2 cell binding could not be observed.

Binding Sites, Antibody↗
PubMed2001

Hemadsorption expressed by cloned H genes from subacute sclerosing panencephalitis (SSPE) viruses and their possible progenitor measles viruses isolated in Osaka, Japan.

Most subacute sclerosing panencephalitis (SSPE) viruses, including our Osaka-1, -2, and -3 strains isolated in Osaka, have shown negative hemadsorption (HAD) by African green monkey red blood cells. This property has been thought to be characteristic of SSPE virus as compared to the positive reaction of the standard Edmonston strain of measles virus (MV). However, this assumption has become quite obscure because MV mutates frequently at the genetic level during its multiplication and also because recent field strains isolated by lymphoblastoid cell lines have shown negative HAD. To investigate the above issue, the nucleotide sequences of the hemagglutinin (H) genes from SSPE virus Osaka-1, -2, or -3 strains were compared to those of various MV field strains isolated in Osaka by Vero cells. The H gene sequences of three SSPE strains were relatively conserved without such biased hypermutation as had been observed in the matrix (M) gene of three SSPE strains. However, this analysis of the H gene sequence of the SSPE viruses enabled us to deduce possible progenitor MVs, which are in agreement with the deduction from the M gene analysis we reported previously. The HAD of Vero cells transfected with the cloned H cDNAs from the SSPE strains and their progenitors suggested that negative HAD of the SSPE viruses has been maintained as one of original properties of the progenitor MVs rather than having been acquired as an altered one during long-term persistent infection in the brains of patients with SSPE.

Amino Acid Sequence↗
PubMed1978

Tissue typing of cells in culture. II. Histocompatibility typing of diploid fibroblastic cultures by the mixed hemadsorption (MH) test.

Skin fibroblasts derived from persons who had been HLA-typed by conventional lymphocytotoxocity tests were subjected to HLA-typing by the mixed hemadsorption (MH) test. The agreement between the two methods was good, provided sera for the MH test were carefully selected. Minor discrepancies found in this study can probably be corrected by further serum selection. There was no difficulty in typing late passages of cultured diploid cells (22 doublings) and in fact it was easier to type a culture at the 33rd doubling than at the 11th. Thus no signs of disappearance of HLA-antigens were seen in these tests.

Cells, Cultured↗
PubMed1971

Production of hemadsorption-negative areas by serums containing Australia antigen.

Exposure of human Wi-38 cells to human serums containing Australia antigen, and presumably serum hepatitis virus, renders the cells refractory to infection by Newcastle disease virus as detected by the hemadsorption-negative plaque test for intrinsic interference. Induction of the Newcastle disease virus refractory state could be passed in cell culture with up to a 1 : 100,000 dilution of material obtained from cells "infected" with serums containing Australia antigen after filtration (0.45-microm pores) and heating to 60 degrees C for 1 hour. Human antiserums to the Australia antigen prevented induction of the Newcastle disease virus refractory state.

Culture Techniques↗
PubMed1974

Microadaption of hemadsorption inhibition for neutralization tests with pig hemagglutinating encephalomyelitis virus.

A microneutralization test is described, in which secondary pig thyroid cells are used and end points are determined by hemadsorption with hamster erythrocytes. The method is evaluated and compared with a macromethod, in which the fluorescent-antibody staining technique is applied for reading titers. The micromethod was found to be useful because of its sensitivity and rapidity.

Analysis of Variance↗
PubMed1976

Use of the hemadsorption phenomenon for determining virus and neutralizing antibody titers of rabies.

Chicken embryo cells infected with the HEP Flury strain of rabies virus adapted to tissue culture produced a hemadsorption (HAD) phenomenon by using goose erthyrocytes. The optimal conditions for HAD included the incubation of cell cultures at 37C for 3 days after virus inoculation, the use of a 0.4% suspension of goose erythrocytes in phosphate buffer adjusted at pH 6.2, and adsorption of erythrocytes at 4C. This phenomenon was inhibited with anti-rabies serum. Virus titer obtained with the HAD technique was almost the same as with the fluorescent antibody technique or the intracerebral inoculation of suckling mice. Results of the neutralization test by using the HAD technique could be easily determined 3 days after inoculation of chicken embryo cells with the mixture of 100 mean tissue culture infective doses of virus and diluted serum. The neutralizing antibody titers coincided with those obtained in mice.

Animals↗
PubMed1982

Identification of Mycoplasma pneumoniae proteins associated with hemadsorption and virulence.

Twenty-two mutants of Mycoplasma pneumoniae spontaneously deficient in hemadsorption were isolated. Examination of mutant protein profiles by one- and two-dimensional polyacrylamide gel electrophoresis permitted the grouping of these mutants into four classes. The largest class of mutants was deficient in four high-molecular-weight proteins (215,000, 210,000, 190,000, and 140,000). A second class of mutants lacked three proteins previously designated A, B, and C (72,000, 85,000, and 37,000, respectively). A single mutant, in addition to lacking proteins A, B, and C, was missing a fourth protein of 165,000 molecular weight. The remaining mutants exhibited protein profiles apparently identical to that of the wild-type strain. All mutant strains attached to the respiratory epithelium of hamster tracheal rings in vitro at reduced levels; however, mutants lacking proteins A, B, and C recognized only neuraminidase-insensitive receptors. None of the mutants tested produced detectable pneumonia in intranasally inoculated hamsters, although one mutant class demonstrated low-level survival in vivo.

Adhesiveness↗
PubMed1983

Inhibition of mycoplasma pneumoniae hemadsorption and adherence to respiratory epithelium by antibodies to a membrane protein.

Antiserum and purified immunoglobulin directed against Mycoplasma pneumoniae membrane protein P1 were examined for their influence on mycoplasma viability, metabolism, and cytadsorption. Anti-P1 immunoglobulin inhibited adherence of M. pneumoniae to hamster tracheal rings in vitro by up to 80% and inhibited hemadsorption by greater than 90%. Cytadsorption was also inhibited by anti-P1 Fab fragments. Anti-P1 antibodies had no effect on M. pneumoniae viability or metabolism. The data indicate that anti-P1 antibody obstructs the interaction of M. pneumoniae adhesin P1 with its host receptors.

Adhesins, Bacterial↗
PubMed2003

Phenotypic switching in Mycoplasma gallisepticum hemadsorption is governed by a high-frequency, reversible point mutation.

Mycoplasma gallisepticum is a flask-shaped organism that commonly induces chronic respiratory disease in chickens and infectious sinusitis in turkeys. Phenotypic switching in M. gallisepticum hemadsorption (HA) was found to correlate with phase variation of the GapA cytadhesin concurrently with that of the CrmA protein, which exhibits cytadhesin-related features and is encoded by a gene located downstream of the gapA gene as part of the same transcription unit. In clones derived from strain R(low), detailed genetic analyses further revealed that on-off switching in GapA expression is governed by a reversible base substitution occurring at the beginning of the gapA structural gene. In HA(-) variants, this event generates a stop codon that results in the premature termination of GapA translation and consequently affects the expression of CrmA. Sequences flanking the mutation spot do not feature any repeated motifs that could account for error-prone mutation via DNA slippage and the exact mechanism underlying this high-frequency mutational event remains to be elucidated. An HA(-) mutant deficient in producing CrmA, mHAD3, was obtained by disrupting the crmA gene by using transposition mutagenesis. Despite a fully functional gapA gene, the amount of GapA detected in this mutant was considerably lower than in HA(+) clonal variants, suggesting that, in absence of CrmA, GapA might be subjected to a higher turnover.

Adhesins, Bacterial↗
PubMed1985

Hemadsorption focus assay for growth of influenza and parainfluenza viruses in human dermal fibroblasts.

An assay for virus-induced hemadsorption foci in cultures of human dermal fibroblasts is described. The assay allows determination of activity of species-specific as well as nonspecific antiviral agents or biological factors, such as interferons. The assay may be used for abortive (e.g., influenza virus) and productive (e.g., parainfluenza virus) infections of human fibroblasts.

Fibroblasts↗