Biomedical subjects

M Zapata

Publications and source records attributed to M Zapata.

At least 19 recordsLinked to original sources

PubMed1998

Cell-fusion assay for the detection of rubella virus in Vero cells.

BACKGROUND AND OBJECTIVES: Rubella virus (RV) produces a subtle and slow-developing cytopathic effect in Vero cells that is difficult to recognize, especially at low multiplicities of infection. In order to facilitate the detection of RV in cell culture, we standardized a low-pH virus-mediated cell-fusion assay. STUDY DESIGN: The incubation periods, temperatures, pH and multiplicity of infection were established. The specificity of the method was tested by immunofluorescence assay and cell-fusion inhibition by specific sera. RESULTS: Six days post infection, Vero cells were treated for 5 min with fusion medium. After that, monolayers were incubated with medium at neutral pH for 16 h and then stained. Gigantic cells with multiple nuclei were observed. CONCLUSIONS: The method allowed the observation of unequivocal images that are easier to recognize than the cytopathic effect caused by RV in the same cell line. At the same time, the method is simple, accessible and shown to be specific to demonstrate the replication of several strains and isolates of RV in Vero cells.

Animals↗

PubMed1995

Separation of mono- and divinyl chlorophyll species by high-performance liquid chromatography using an octadecyl polyvinyl alcohol polymer column.

Most of the chlorophylls and their related compounds from nonesterified chlorophyllide to esterified pheophytin were separated by high- performance liquid chromatography using a wide pore, C(18) polyvinyl alcohol polymer column with an elution using a binary gradient starting from a buffered mobile phase. The high selectivity of this system enabled not only the separation of common chlorophyll species but also the resolution of structurally similar compounds such as mono- and divinyl chlorophyllide and pheophorbide species which usually coelute on monomeric bonded phases. The method is successfully applied to analyses of the pigments extracted from green and etiolated tissues of higher plants and photosynthetic bacterial cells.

Chlorophyll↗

PubMed1995

Rapid HCV RNA detection by PCR followed by a new non-radioactive liquid hybridisation assay and comparison with RIBA.

A one-stage polymerase chain reaction (PCR) followed by an automated liquid hybridisation assay was used to examine anti-HCV-positive patients. The presence of HCV-RNA in 251 randomly selected enzyme immunoassay (EIA) positive clinical specimens was compared to their RIBA pattern. An association of a RIBA pattern with presence or absence of HCV-RNA was not detected. One hundred of these samples were also evaluated after dividing them into normal and elevated serum alanin aminotransferase (ALT) levels. PCR results were obtained on the basis of amplification products from two different gene regions (5'-NC region and NS 3). To prove the specificity of the PCR products, a commercially available digoxigenin-based liquid hybridisation assay was evaluated. The sensitivity was comparable to the results obtained after nested PCR. Based on the results of the study, the two-stage PCR can be changed in favour of the easier one-step PCR which offers the advantage of fewer contamination problems.

Alanine Transaminase↗

PubMed1991

Kinetics of rubella-specific IgM antibody response in postnatal rubella infection.

The serological diagnosis of primary postnatal rubella infection is based on detection of rubella-virus-specific IgM antibody or a four-fold rise in rubella-specific IgG antibody. Although there are several different methods of enzyme immunoassays that are commercially available, the cost benefit evaluation makes them impractical for use in developing countries. For this reason, we have standardized the measurement of rubella IgM antibody by HAI following serum fractionation by ion-exchange chromatography. The sera samples obtained from pregnant women infected with rubella virus at different times during gestation were fractionated and tested by HAI. Seven out of nine sera collected within the first two days after onset of rash showed detectable levels of rubella IgM antibody. All 57 sera collected between 3 and 30 days after the onset of rash contained rubella IgM antibody. After 30 days, only 1 of 5, or 20%, of sera contained IgM antibody. The HAI testing method was rapid and specific and the cost was not prohibitive. HAI-IgM testing could be used to diagnose primary rubella infections in developing countries where expensive EIAs are unaffordable.

Antibodies, Viral↗

PubMed1989

Comparison of mitogenic responses of young and old rhesus monkey T cells to lectins and interleukins 2 and 4.

Purified T cells from rhesus monkeys, like human T cells, do not show a significant mitogenic response to lectins or PMA, but when combined with PMA or accessory cells, PHA and Con A induce a vigorous mitogenic response. This response is strongly impaired in purified T cells from old rhesus monkeys compared to young T cells, from 56 to 72%, and parallels results obtained with T cell preparations containing accessory cells. Likewise, purified T cells do not respond to interleukin 2 (IL-2) or IL-4, but in the presence of PMA, a significant mitogenic response occurs in the young but not the old T cells. This response is augmented by accessory cells, but is still very deficient in the old T cells. These results show that the IL-2 independent activation of T cells triggered by IL-4, like the conventional IL-2 activation, is age impaired. The deficient response to IL-2 implies an age-related deficiency in IL-2 receptor as well in aged rhesus T cells, and may account for the less effective response of the old cells to calcium ionophore (+PMA) activation. The use of purified T cells in these studies obviate the influence of accessory cells, and thus simplify interpretation.

Age Factors↗

PubMed1989

Comparative mitogenic response of old and young rhesus monkey T cells to lectin from Erythrina cristagalli.

The lectin (EC) from the coral tree, E. cristagalli, while less mitogenic on a molar or weight basis than PHA or ConA, strongly activates both Rhesus monkey and human T cells. The optimal mitogenic concentrations for both Rhesus and human T cells are 0.25, 2.5, and 25 micrograms/ml, respectively, for PHA, ConA, and EC. Aged Rhesus T cells were profoundly suppressed in mitogenic response to EC (approx. 80%) compared to young Rhesus cells. However, in the presence of supplemental interleukin 2 (20 U/ml), the age-related defect was reversed; the average mitogenic response of the old Rhesus T cells was increased sixfold.

Aging↗

PubMed1989

Characteristics of different solid-phase immunoassay formats for the measurement of BK virus immunoglobulin M in sera of patients on renal dialysis or with kidney allografts.

Solid-phase immunoglobulin M (IgM) antigen capture enzyme immunoassay (AgCEIA) and antibody capture enzyme immunoassay (AbCEIA) were developed for the diagnosis of BK virus (BKV) infections. Of 37 serum samples from renal allograft recipients, 15 were positive for BKV IgM antibody by either AgCEIA, AbCEIA, or antigen capture radioimmunoassay. False-positive IgM results were observed in the AgCEIA in the presence of high levels of BKV IgG antibody (titers greater than or equal to 1:51,200), when rheumatoid factor (RF) titers were greater than or equal to 1:20, or in the presence of high levels of RF (titers greater than or equal to 1:10,240) when BKV hemagglutination inhibition titers exceeded 1:40. False-positives due to RF could be eliminated by treatment of sera with anti-human IgG antisera or IgG-coated latex particles. The presence of RF did not, however, produce false-positive results in the AbCEIA. Both AgCEIA and AbCEIA were specific for BKV IgM antibody, as 14 serum samples containing either JC papovavirus, cytomegalovirus, rubella virus, hepatitis A virus, or hepatitis B virus core IgM antibody were negative in both EIAs. Comparison of results obtained for 37 serum samples revealed 14 positive by radioimmunoassay and 11 positive by both AgCEIA and AbCEIA. Both EIAs detected BKV IgM antibody in sera of renal allograft patients and patients on renal dialysis who had reactivated BKV infections persisting for several months after transplantation.

Antibodies, Viral↗

PubMed1988

Excess incidence of treatment of end-stage renal disease in Mexican Americans.

Mexican Americans are the second largest minority group in the United States (8.73 million people according to the 1980 US census) and are known to have an excess prevalence of obesity and non-insulin-dependent diabetes mellitus, but similar or lower rates of hypertension when compared with non-Hispanic whites. To our knowledge, no data are available on incidence of end-stage renal disease in this population. Using a data base from the Texas Kidney Health Program, a division of the Texas Department of Health, and the 1980 US census for the state of Texas, the authors calculated age-adjusted incidence of treatment of end-stage renal disease in Mexican Americans, non-Hispanic whites, and blacks for the years 1978-1984. Mexican Americans and blacks have an excess of treatment of end-stage renal disease (all etiologies combined) compared with non-Hispanic whites (incidence ratios of 3 and 4, respectively). For diabetes-related end-stage renal disease, Mexican Americans have an incidence ratio of 6, while blacks have an incidence ratio of 4 compared with non-Hispanic whites. For Mexican Americans, this excess is higher than would be expected on the basis of their underlying prevalence of diabetes. The incidence of hypertensive end-stage renal disease in Mexican Americans was 2.5 times higher than in non-Hispanic whites, which is higher than expected given the lack of excess in their underlying prevalence of hypertension. The high prevalence of diabetes in Mexican Americans explains some, but not all, of the excess of treatment of end-stage renal disease in this population.

Adult↗

PubMed1986

[Morphologic characteristics of plaques produced by the rubella virus in cultures of Vero cells].

Under defined plaquing condition, Gilschrist strain of rubella virus showed different plaque morphology and plaque size when tested in Vero cell cultures. These differences were obtained by changing the fetal bovine serum (FBS) concentration. With 2 per cent of FBS rubella virus formed clear plaques that included a number of cells which retained the stain, while with 4 per cent of FBS it formed ring-shaped plaques. These characteristics were retained even after the Gilschrist strain was passaged several times in Vero cell cultures. The cytopathic effect in Vero cell cultures proved to be useful for the titration of rubella virus, giving infective titres in the same logarithmic order than the plaque assay.

Animals↗

PubMed1984

Measurement of BK papovavirus IgG and IgM by radioimmunoassay (RIA).

Current techniques for the measurement of BK papovavirus (BKV) specific IgM include sucrose density gradient centrifugation followed by hemagglutination inhibition (HAI) or indirect immunofluorescent (IF) staining of BKV infected cells using a fluorescein conjugated anti-human IgM antibody. These techniques are cumbersome and labor intensive and do not lend themselves to testing large numbers of sera. A solid phase radioimmunoassay (RIA) was developed to facilitate the measurement of BKV IgG and IgM in large numbers of sera. Solid phase antigen was prepared by adsorbing CsCl purified BKV antigen to polyvinyl chloride microtiter plates. Following reaction with serum, bound immunoglobulin was detected with iodinated goat anti-human IgG or IgM. RIA for the measurement of BKV IgG was sensitive with titers approaching 10(-6). Determination of IgG titers by RIA and HAI showed good agreement (P less than 0.01, correlation coefficient = 0.74). Measurement of BKV IgM was not affected by the presence of BKV IgG as evidenced by sucrose density gradient fractionation of IgM positive sera, removal of IgG by treatment with S. aureus protein A, and addition of BKV IgG to BKV IgM. Rheumatoid factor (RF) gave false positive IgM titers in the presence of BKV IgG when RF titers were greater than or equal to 1:640 by latex agglutination testing and BKV IgG levels exceed 1:256 by HAI. False positives due to RF could be eliminated by treatment of sera with sheep anti-human IgG antisera. RIA for BKV IgM was specific as sera containing JCV-, cytomegalovirus (CMV)-, rubella-, or hepatitis B core antibody (anti HBc)-IgM were negative by RIA. RIA detected BKV IgM in several sera from renal dialysis or allograft patients with titers ranging from 1:400 to 1:128,000 and demonstrated that BKV IgM persisted in sera of renal allograft patients for as long as 343 days post transplantation.

Antibodies, Viral↗

PubMed1984

Indirect immunofluorescence staining of Chlamydia trachomatis inclusions in microculture plates with monoclonal antibodies.

Indirect immunofluorescence (IF) staining, using a monoclonal antibody, detected two- to fourfold more inclusions than did iodine staining. Of 274 clinical specimens, 53 (19.3%) were positive by IF on passage 1 as compared with 33 (12%) by iodine staining (P less than 0.005). IF-stained inclusions in McCoy cells in the bottom of microculture wells were readily viewed with a long-focal-length objective at a magnification of 250 X.

Antibodies, Bacterial↗

PubMed1974