Dot immunoperoxidase assay using monoclonal antibody for direct detection of cytomegalovirus in urine samples

Zerbini, Marialuisa; Musiani, Monica; Gentilomi, Giovanna Angela; Plaça, M. La

doi:10.1128/jcm.25.11.2197-2199.1987

Cited by 16 publications

(4 citation statements)

References 18 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Because nitrocellulose sheets (NCS) strongly adsorb proteins and nucleic acids, they are widely used for various biological assays, including the detection of viral genomes and antigens from patient specimens (1,3,9,16) and the detection of antiviral specific antibodies in patient serum by reaction with antigens immobilized on the NCS (4,5). In the present study, we tried to apply this technique to the serodiagnosis of scrub typhus.…”

mentioning

confidence: 99%

Serodiagnosis of scrub typhus with antigens immobilized on nitrocellulose sheet

et al. 1989

View full text Add to dashboard Cite

This study was designed to develop a simple method for serodiagnosis of scrub typhus. The basis of the method is detection of anti-Rickettsia tsutsugamushi antibody in patient serum by reaction with antigens dot blotted on a nitrocellulose sheet (NCS). The final evaluation of the reaction is performed by observing the color intensity which develops as a result of sequential treatments of the NCS with peroxidase-conjugated anti-human immunoglobulin G or immunoglobulin M antibody and with the substrate of the enzyme. After various trials, we found that the best results were obtained by using a purified antigen which adhered to an NCS at 0.2 to 2 ,ig of protein per dot and a test serum diluted 1,000to 4,000-fold. Under these conditions, almost all antibody-positive sera showed a distinct color at the dot on the NCS, so that a positive reaction could be distinguished by the naked eye from a negative reaction with antibody-negative sera, which developed only a faint color. A comparison of the results of screening of antibody-positive and-negative sera by this method and the immunofluorescence test showed that both methods produced similar results. From these results, it is concluded that this dot immunoassay can be useful for the serodiagnosis of scrub typhus.

show abstract

mentioning

confidence: 99%

Serodiagnosis of scrub typhus with antigens immobilized on nitrocellulose sheet

et al. 1989

View full text Add to dashboard Cite

show abstract

“…In our assay, serum specimens were treated with SDS denaturing buffer in order to expose the antigenic sites present in the virions and to avoid the masking of antigenic determinants due to the presence of host-specific antibody (19). This treatment was also able to inactivate up to 500 g of horseradish peroxidase per ml, thus avoiding any possibility of cross-reactivity due to endogenous peroxidase present in serum specimens.…”

Section: Discussionmentioning

confidence: 99%

Dot immunoperoxidase assay for detection of parvovirus B19 antigens in serum samples

et al. 1997

Self Cite

View full text Add to dashboard Cite

We describe a simple and rapid dot immunoperoxidase assay for the direct detection of parvovirus B19 capsid antigens in human sera. The assay was performed with serum specimens dotted onto nylon membranes. VP1 and VP2 B19 antigens, which represent 4 and 96% of the capsid, respectively, were detected with a pool of monoclonal antibodies directed against the two proteins, and the complex was visualized by immunoperoxidase staining. The assay could be performed in about 4 h, and positive results were revealed at the end of the reaction as dark blue spots on the nylon membrane at the site of positive specimens. A total of 541 serum samples from different subjects and with different laboratory evaluations with regard to B19 infection were analyzed. The results obtained by the dot immunoperoxidase assay were compared with the results obtained for the presence of B19 DNA by dot blot hybridization and nested PCR. With optimized working conditions, the dot immunoperoxidase assay was able to detect the presence of B19 with a sensitivity comparable or slightly higher than that achieved by dot blot hybridization but less than that achieved by nested PCR. Since the level of sensitivity of the dot immunoperoxidase assay proved to be appropriate for the detection of acute B19 infection, and since the cost, time to a result, and versatility of the assay are important issues, from our evaluation, the dot immunoperoxidase assay described may be particularly suitable for large-scale screening of samples and a good alternative to DNA detection methods in the routine laboratory evaluation of B19 infection.

show abstract

“…However, tissue culture is expensive and time-consuming and requires prompt processing of the specimens in a tissue culture laboratory. Alternative procedures, such as the detection of viral antigens or viral DNA, have recently been developed (2)(3)(4)(5)(6)(7), but one of the major disadvantages of these techniques is the need to ultracentrifuge the urine in order to have highly Concentrated viral material (2)(3)(4)(5)(6)(7). This consideration prompted us to test several different procedures to concentrate CMV from urine without ultracentrifugation.…”

Section: Introductionmentioning

confidence: 99%

A simple and rapid procedure for the direct detection of cytomegalovirus in urine samples

Landini

Guan

Ripalti

et al. 1990

Clinical Laboratory Analysis

View full text Add to dashboard Cite

We compared cytomegalovirus (CMV) isolation in tissue culture and viral DNA detection by DNA:DNA hybridization in 60 clinical urine samples concentrated by different procedures. Our results showed that urine concentration by filtration is the easiest and quickest procedure allowing further detection of CMV. With optimized working conditions, the CMV detection in urine can be done in 5 hr without requiring cell cultures or costly instruments.

show abstract

Dot immunoperoxidase assay using monoclonal antibody for direct detection of cytomegalovirus in urine samples

Cited by 16 publications

References 18 publications

Serodiagnosis of scrub typhus with antigens immobilized on nitrocellulose sheet

Serodiagnosis of scrub typhus with antigens immobilized on nitrocellulose sheet

Dot immunoperoxidase assay for detection of parvovirus B19 antigens in serum samples

A simple and rapid procedure for the direct detection of cytomegalovirus in urine samples

Contact Info

Product

Resources

About