Demonstration of Rubella IgM Antibody by Indirect Fluorescent Antibody Staining, Sucrose Density Gradient Centrifugation and Mercaptoethanol Reduction

Forghani, Bagher; Schmidt, Nathalie J.; Lennette, Edwin H.

doi:10.1159/000148832

Cited by 38 publications

(22 citation statements)

References 12 publications

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“…In the present study, all results of the IPA test performed on whole serum were confirmed on serum fractions. Problems of nonspecific staining produced by low dilutions of some human sera, as reported using the I FA technique for rubella IgM detection [18]. did not interfere critically with the reading of test results.…”

Section: Discussionmentioning

confidence: 84%

Rapid Detection of Human Cytomegalovirus and Herpesvirus Hominis IgM Antibody by the Immunoperoxidase Technique

Gerna

Chambers

1977

Intervirology

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Immunoglobulin M (IgM) to human cytomegalovirus (CMV) and herpesvirus hominis (HVH, types 1 and 2) has been searched for in a group of patients with congenital and recent CMV and HVH infections and also in a group of patients with remote CMV and recurrent HVH infections, using the immunoperoxidase antibody technique (IPA) as compared to a technique using sucrose density gradient differential centrifugation. Both techniques detected the same positive samples. High IgM antibody titers (1:32–1:512) were observed in all cases of primary CMV and HVH infection, whereas low titers (1:8–1:16) were found in congenital infections. The IPA technique is sensitive, specific and useful for rapid diagnosis of recent and congenital CMV and HVH infections.

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Section: Discussionmentioning

confidence: 84%

Rapid Detection of Human Cytomegalovirus and Herpesvirus Hominis IgM Antibody by the Immunoperoxidase Technique

Gerna

Chambers

1977

Intervirology

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“…This method was criticized by Newman et al (1969), who found, when examining cord sera from infants with congenital rubella, that the IgM fractions were consistently contaminated with IgG which they thought might contribute to the HAI activity. Forghani, Schmidt & Lennette (1973) also detected IgG in the IgM fractions from some adult sera and suggested that this might be due to aggregation of IgG. Caul, Smyth & Clarke (1974) attempted to exclude false positive results due to contamination with IgG by titrating the HAI activity before and after treatment with 2-mercaptoethanol (2ME) which reduces IgM.…”

Section: Discussionmentioning

confidence: 99%

Specific immunoglobulins in infants with the congenital rubella syndrome

Cradock-Watson

Ridehalgh

1976

J. Hyg.

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SUMMARYThe indirect immunofluorescent technique has been used to detect and titrate the specific immunoglobulins in serum specimens from 154 infants with confirmed or suspected congenital rubella. IgM antibody was stained more efficiently in sucrose density gradient fractions than in whole serum and was detected in this way in 27 out of 40 patients with confirmed congenital rubella at ages ranging from birth to 2 years. It was present in 48 out of 50 serum specimens during the first 6 months of life and in 11 out of 38 specimens obtained at ages between 62 months and 2 years. IgM antibody was therefore estimated to persist for about 6 months in the majority of cases and up to 2 years in a few individuals. IgM antibody was also detected by this method in 11 out of 114 infants with suspected but unconfirmed congenital rubella at ages up to 5 months. The total concentrations of IgM were above the normal range in nearly all sera taken from confirmed cases during the first 3 months of life and in half the specimens obtained between the ages of 3 and 6 months.IgG antibody was detected by fluorescent staining of whole serum in all patients with congenital rubella. Geometric mean titres increased during the first 3 months of life and then declined slowly. IgA antibody was not detected, except in two patients in whom traces were present at the age of 6 months, and the total concentrations of IgA were usually within normal limits.Fluorescent staining of fractions showed that the sedimentation characteristics of rubella IgG and IgM antibodies were the same in infants as in adults. The peak IgM fractions never contained IgG antibody, and the presence of specific IgM in these fractions could usually have been safely inferred from their IIA titres. Fluorescent staining, however, was more sensitive and frequently detected IgM antibody in fractions which had no definite HAI activity.Fluorescent staining of whole serum for IgM antibody was less distinct, and often unsuccessful, even in specimens in which specific IgM was detected in the fractions. The addition of IgG-to IgM-containing fractions caused depression of IgM staining and suggested that failure to detect IgM antibody in whole serum was partly due to competitive inhibition by specific IgG.

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“…The test was simplified by Desmyter et al (1971), who examined two fractions but detected no IgM after 36 days. Forghani et al (1973) also tested only two fractions, and found specific IgM in all of 18 sera taken between 3 and 20 days after onset, and none of 10 subjects taken 1-30 years after rubella. A simplified test with only one fraction tested with and without 2ME similar to the one described here was developed by Field & Murphy (1972).…”

Section: Discussionmentioning

confidence: 99%

“…Both trailing and mixing are controlled by treating the fractions with 2ME, which shows that the inhibition is due to 337 IgM and not IgG. Forghani et al (1973) suggested that 2ME is not satisfactory for confirming IgM in fractions because reaggregation may take place, but Field & Murphy and ourselves have found no difficulty in confirming IgM in the fractions with 2ME.…”

mentioning

confidence: 90%

“…Conventional laboratory tests may not help to decide whether the patient has had rubella during her pregnancy. The presence of rubella-specific IgM immunoglobulin seems to indicate recent infection (Vesikari & Vaheri, 1968;Best, Banatvala & Watson, 1969;Desmyter, South & Rawls, 1971;Ogra et al 1971; Cradock-Watson, Bourne & Vandervelde, 1972;Haire & Hadden, 1972;Field & Murphy, 1972;Forghani, Schmidt & Lennette, 1973). This has been detected by 2-mercaptoethanol (2ME) treatment of whole serum (Banatvala et al 1967), immunofluorescence (Baublis & Brown, 1968; Cohen, Ducharme, Carpenter & Deibel, 1968), separation of immunoglobulins in sucrose density gradients followed by rubella haemagglutination inhibition (HAI) (Vesikari & Vaheri, 1968) or radioimmunodiffusion (Ogra et al 1971), separation in a Sephadex column (Gupta, Peterson, Stout & Murphy, 1971) or agarose (Burgin-Wolff, Hernandez & Just, 1971) followed by HAI.…”

Section: Introductionmentioning

confidence: 99%

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A simplified method for the detection of rubella-specific IgM employing sucrose density fractionation and 2-mercaptoethanol

Caul¹,

Smyth²,

Clarke³

1974

J. Hyg.

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SUMMARYA simplified method for the detection of rubella-specific IgM in sera was developed involving HAI tests on only one fraction from a sucrose density gradient with and without treatment with 2-mercaptoethanol (2 ME). Slight trailing of IgG into the IgM fraction was shown to occur in patients with high titres of both IgG and IgM, but this could be detected by the 2ME treatment and did not affect the results.By this test, rubella-specific IgM was found in all of 54 patients with a rash and a rise of rubella HAI antibodies. When the antigen and serum were incubated for 1 hr. before adding the RBC, specific IgM was found in all 51 sera taken between 5 and 40 days after onset, and in about half the sera taken between 42 and 77 days, but in none of 16 people known to have possessed rubella antibodies at least a year previously. When antigen and serum were incubated for 18 hr. before adding the RBC, specific IgM was found in all of 21 sera taken between 2 and 49 days after onset, and in 6 of 11 sera taken between 50 and 117 days, but not in 9 people known to have possessed rubella antibodies at least a year before. The method has been found to be very useful in detecting recent rubella among patients in early pregnancy.

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Demonstration of Rubella IgM Antibody by Indirect Fluorescent Antibody Staining, Sucrose Density Gradient Centrifugation and Mercaptoethanol Reduction

Cited by 38 publications

References 12 publications

Rapid Detection of Human Cytomegalovirus and Herpesvirus Hominis IgM Antibody by the Immunoperoxidase Technique

Rapid Detection of Human Cytomegalovirus and Herpesvirus Hominis IgM Antibody by the Immunoperoxidase Technique

Specific immunoglobulins in infants with the congenital rubella syndrome

A simplified method for the detection of rubella-specific IgM employing sucrose density fractionation and 2-mercaptoethanol

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