Virus-specific IgM Antibodies Research Articles

Human cytomegalovirus infection: recent developments in diagnosis and epidemiology.

Cytomegalic inclusion disease (CID) is caused by a horizontally or vertically transmitted human herpes virus infection and may persist for life without obvious clinical symptoms. A serious course of horizontal primary and recurrent infections, however, is often observed in immunocompromised persons such as recipients of organ transplants and patients receiving fresh blood transfusions. Vertical infection may cause fetopathies. The human cytomegalovirus (HCMV) is thought to inherit an oncogenic potential as lately discussed for AIDS and M. Kaposi. Laboratory diagnosis of HCMV infection is performed by light microscopy (inclusion bodies), electron microscopy, virus isolation in cell culture, demonstration of viral DNA and antigen in clinical specimens, by histochemical methods (e.g. immunoperoxidase technique) and by DNA and peptide analysis for identification of different isolates and viral finger prints. Evaluation of cell-mediated immunity in HCMV infection is performed quantitatively (assessment of Thelper/Tsuppressor ratios) or qualitatively (specific lymphocyte stimulation by the antigen). In most cases laboratory diagnosis is achieved by serological methods, i.e. demonstration and quantitation of HCMV-specific antibodies. In this context, a number of liquid- and solid-phase immunoassays have been developed, of which immunofluorescence and ELISA are most commonly used, besides complement fixation and passive haemagglutination. These procedures on the one hand allow the use of different antigen preparations as early and late viral proteins, and on the other hand permit a specific determination of different Ig classes and subclasses. A variety of assays has been established especially for determination of virus-specific IgM antibodies, which are predominantly found in active infection. These, however, at least in part may show non-specific results caused by interference of rheumatoid factor or IgG competition. Such problems have now been dealt with and are avoided by IgG precipitation or IgM immunosorption ("mu-capture" technique). These recent methods allow an exact epidemiological identification of risk groups for CMV infection. Results from our laboratory revealed 13% HCMV-IgM positive patients among pregnant women, 16% IgM positive patients among renal transplant recipients, 4% IgM positive cases in patients after cardiosurgery and 1.7% IgM positives among prostitutes. The prevalence of HCMV infection as indicated by specific IgG antibodies was 56%, 90%, 83%, and 90%, respectively.(ABSTRACT TRUNCATED AT 400 WORDS)

DETECTION OF MEASLES IgM

Fewer physicians have experience in diagnosis of measles as the number of cases continues to decline. For this reason an Enzyme Linked Immunosorbent Assay (ELISA) was developed to detect measles virus specific IgM antibody (MIgMA). Anti-human μ chain is affixed to a solid phase to which a 1:100 dilution of serum is added. Only 0.005 ml of patient's serum is needed. Treatment of 3 MIgMA positive sera with dithiothreitol but not Staph protein A removed MIgMA. The values for 24 cord sera (0.030 ± 0.007), 60 adult sera (0.034 ± 0.011), and 47 sera from children prior to measles immunization (0.030 ± 0.015) were used to establish a seronegative range. Sera yielding reactions greater than 3 SD of the mean for the latter group were considered to be positive for the presence of MIgMA. MIgMA was not detected in sera with high titers of rheumatoid factor. Five unpaired and the first of 14 of 17 paired sera obtained from patients with measles were positive for MIgMA. The 3 convalescent samples from patients having MIgMA negative initial sera, also were positive. MIgMA was detected as early as 1 day and as late as 41 days following onset of illness. No MIgMA could be detected in 47 vaccinees tested 2-1/2 months or more after measles vaccination. Neither recent immunization nor the presence of rheumatoid factor produce reactions which are likely to obfuscate the interpretation of this very sensitive test which, in most cases, can confirm the clinical diagnosis of measles on a single serum specimen obtained soon after onset of rash.

Immunoglobulin classes of Aleutian disease virus antibody.

Aleutian disease virus (ADV) persistently infects mink and causes marked hypergammaglobulinemia. Immunoglobulin class-specific antisera were used to define the total immunoglobulin of each class by radial immunodiffusion and the immunoglobulin class of ADV-specific antibody by immunofluorescence in experimentally and naturally infected mink. Electrophoretic gamma globulin closely reflects the immunoglobulin G (IgG) level in mink, and the majority of the increased immunoglobulin and ADV antibody in infected mink is IgG. IgM becomes elevated within 6 days after infection, reaches peak levels by 15 to 18 days, and returns to normal by 60 days after infection. The first ADV antibody demonstrable is IgM, and most mink have virus-specific IgM antibody for at least 85 days postinfection. Serum IgA levels in normal mink are not normally distributed, and ADV infection causes a marked elevation of IgA. Low levels of ADV-specific IgA antibody can be shown throughout the course of infection. Failure of large amounts of virus-specific IgG antibody to inhibit the reaction of virus-specific IgM and IgA antibodies suggests that the various classes of antibodies are directed against spatially different antigenic determinants. The IgM and IgA were shown not to be rheumatoid factors.

Hepatitis A virus immunoglobulin M antibody in acute neurological disease.

Sera from 167 patients with Guillain-Barré syndrome, 224 patients with other neurological diseases, and 10 normal subjects were tested for hepatitis A virus-specific immunoglobulin M antibodies. Two patients with classic Guillain-Barré syndrome and 1 patient with acute myeloradiculopathy had high titers of virus-specific IgM antibody. All three patients had jaundice and laboratory evidence of liver involvement before the onset of the neurological disease. None of the other sera from patients with Guillain-Barré syndrome or control subjects were positive for hepatitis A virus.

Rapid Viral Diagnosis

There has been much interest and activity in the development of techniques for rapid viral diagnosis which would allow successful intervention in the treatment of patients or their contacts or in the control of viral diseases in the community. The greatest emphasis has been on techniques that permit viral detection directly in the clinical specimen, since these avoid the need to cultivate the agent, are feasible for detection of viruses that cannot be cultivated, and can detect virus in specimens in which the agent is no longer infectious. Direct methods used for viral detection include electron microscopy and various immunoassays which are based on demonstrating reactivity of viral antigen in the specimen with known viral antisera. The use of immunoassays for more rapid identification of viruses isolated in laboratory host systems and for selective detection of viral antigen in inoculated cell cultures even before the agents produce an observable effect has been an important advance in viral diagnosis by the approach of isolation and identification. The reliability of all specific viral identification procedures depends on the use of high quality viral antisera. Some of the problems previously encountered in preparing satisfactory viral immune reagents are being overcome through the availability of highly specific monoclonal antibodies produced by cell hybridization techniques. Virus-specific IgM antibody assays for rapid diagnosis have been improved greatly through the use of a "capture" technique in which antibody to the human mu chain is used in the solid phase to separate IgM from other serum components which might compete with IgM antibody or give nonspecific reactivity, and also through the availability of highly specific monoclonal antibodies to the human mu chain. A variety of simple assays for determination of viral antibody status have been developed, and many are commercially available. The reliability of some of these antibody assays has been improved through the incorporation of more suitable controls and through better definition of interpretations which should be made from test results.

Measles virus-specific IgM antibodies in sera from patients with chronic active hepatitis.

The occurrence of measles virus-specific IgM antibodies in sera from patients with chronic active hepatitis not caused by Hepatitis B virus was examined by a specific enzyme-linked immunosorbent assay (ELISA). Using whole serum, specific IgM antibodies were detected in 12 of 23 sera from patients with hepatitis B surface antigen (HBsAg)-negative chronic active hepatitis. In nine of these sera the finding of specific IgM antibodies was verified by separation of IgM by density gradient centrifugation and examination of the fractions by ELISA. Most of the sera from the patients with measles virus-specific IgM antibodies had an elevated level of specific IgG antibodies compared to the level of IgG found in control sera. The significance of these findings in view of a possible persistent measles virus antigen production in patients with chronic active hepatitis is discussed.

Cerebrospinal fluid IgG bands and virus-specific IgG, IgM, and IgA antibodies in herpes simplex virus encephalitis

To characterize the immune response of the central nervous system in herpes simplex virus (HSV) encephalitis, cerebrospinal fluid (CSF) specimens of 7 biopsy proven and 7 presumptive herpes simplex virus (HSV) encephalitis patients were studied, using sodium dodecyl sulfate polyacrylamide gel electrophoresis for the presence of CSF IgG bands, and solid-phase enzyme immunoassays for HSV-specific antibodies. IgG bands were detected in all CSF specimens of the patients, as early as day 6 and up to day 1088. A novel, unidentified, 120 000 dalton polypeptide was found in the CSF of most of the patients, in a total of 2550 specimens, but not in the controls. This polypeptide was evident by day 6, its intensity fluctuated and it was present in specimens collected as late as day 855. HSV-specific antibodies, of either IgG, IgM, or IgA class, were not detected in the CSF during the first week of illness. IgG antibodies appeared later in all patients and persisted to the end of the follow-up of 3 years. The fact that CSF IgG bands were present in some patients before the appearance of HSV antibodies, and also persisted longer, suggests that the IgG response is not restricted to HSV-specific antigenic determinants.

Intrathecal synthesis of virus-specific oligoclonal IgG, IgA and IgM antibodies in a case of varicella-zoster meningoencephalitis

Varicella-zoster (VZ) virus meningoencephalitis was diagnosed in a 72-year-old man without other clinical signs of VZ infection, on the basis of intrathecal virus-specific IgG, IgA and IgM antibody responses demonstrated by imprint immunofixation (IIF) and by serological analyses of serum and CSF. The intrathecally produced antibodies displayed oligoclonal characteristics. The intrathecal production of VZ-IgG and -IgA antibodies persisted throughout the observation period of 20 months, while that of VZ-IgM antibodies was not detectable later than 3 months after onset. Part of the intrathecally produced VZ-IgG and -IgA antibody populations, but no IgM antibodies, were shown to cross-react with herpes simplex virus. Oligoclonal IgG bands were demonstrated in the CSF throughout the observation period. The bulk of the IgG bands was shown to represent VZ-specific antibodies.

Virus-specific IgG and IgM antibodies in normal and immunocompromised subjects infected with cytomegalovirus.

Levels of IgG and IgM antibodies t human cytomegalovirus (CMV) were measured using a solid-phase radioimmunoassay. Individuals positive by complement-fixation test consistently had detectable IgG titers by radioimmunoassay, but no quantitative relationship was apparent. An elevated IgM titer was considered specific for CMV infection because sera from individuals with other herpesvirus infections did not cross-react. In patients with mononucleosis, elevated titers of IgM antibody to CMV correlated (P less than 0.001) with active infection and were highest during viremia. Titers of IgG antibody to CMV during and after symptomatic infection were similar to those of asymptomatic positive individuals. Increases in CMV-specific IGM were observed in both primary and reactivated infections in cardiac transplant recipients. In a small group of cardiac transplant recipients with recurrent symptomatic disease, IgM titers were low at the time of viruria and did not increase with CMV tissue involvement, a result which suggests that quantitative deficiencies in IgM may be related to the severity of CMV infections.

Beta 2-microglobulin serum levels in infectious mononucleosis in childhood.

A material of fifteen children aged 10 months to 15 years with infectious mononucleosis (IM) was investigated. The diagnosis was established by demonstration of Epstein-Barr virus (EBV) specific IgM antibodies. The serum concentrations of beta 2-microglobulin (beta 2-m) were significantly increased (p less than 0.01) in the first sample after onset of disease, compared to control groups of children with other infectious diseases and children without diseases. The serum concentration of beta 2-m decreased towards normal range within 3 weeks- 3 months. During the same period, IgM antibodies of EBV decreased to unmeasurable levels, and the IgG titers were almost constant. Serum beta -2m may be a parameter of value in the diagnosis of infectious mononucleosis in childhood, replacing tests for heterophile antibodies (Emni test and Paul-Bunnell test) which are often negative in children suffering from a primary EBV infection, or it may be used for assessment of disease activity.

A modified absorption-reduction method to detect virus-specific hemagglutination inhibiting and neutralizing IgM antibodies.

IgG antibodies are preferentially absorbed by protein A-coupled sepharose beads while most of the IgM and IgA antibodies remain in solution. Even relatively low amounts of virus-specific IgM antibodies can then be detected unequivocally by a decrease of the antibody titer following the treatment with reducing agents. Ethandithiol proved superior to 2-mercaptoethanol in combination with neutralization assays.

Anticorps inhibant l'hémagglutination du virus BK: Brève surveillance de 52 transplantés rénaux et prévalence dans différents groupes d'âge de l'ouest de la France

Summary Screening of haemagglutination-inhibiting antibodies of a BK virus (Gardner strain) for purposes of sero-epidemiological follow-up was performed on sequential sera obtained from 52 kidney allograft recipients during the first three months following grafting as well as on serum obtained from the same subjects before grafting. Thirty-nine recipients (75%) had anti-BK-virus antibodies before grafting, 13 of whom exhibited viral reactivation with a significant rise in antibodies. Only one patient had seroconversion, indicating primary viral infection. Fractionated sera from 6 subjects were screened for specific IgM BK-virus antibodies which were found not only in the first positive serum from primary infection, but also in 5 reactivation sera. The customary method of testing for virus-specific IgM antibodies was used: screening for the rheumatoid factor, sensitivity to 2-mercapto-ethanol and verification of the separation procedure by radial immunodiffusion. No correlation with clinical symptoms or changes in renal function was noted in subjects with a rise in BK-virus antibodies. In 3 cases, rises in anti-BK-virus antibodies coincided with a rise in anti-cytomegalovirus antibodies. These results were compared with those of blood samples from 50 healthy adult donors and 133 children ranging from 1 month to 14 years of age, living in the same region (western France), as the kidney allograft recipients. Adults tested 82% positive for BK-virus antibodies, and children over 4 years of age 69%, which indicates early acquisition of the antibody in question.

Serologic studies on the association of rubella virus infection and juvenile rheumatoid arthritis.

Rubella haemagglutination-inhibition (HAI) and virus-specific IgM antibody responses were investigated in 26 patients with Juvenile Rheumatoid Arthritis (JRA) and 52 age-and sex-matched controls. 30.8% of patients with JRA showed no serological evidence of a previous infection by rubella virus (RV). In addition, the Geometric Mean Titres (GMT) of rubella HAI antibody among patients with JRA were similar to those found among the control group. Furthermore, rubella-specific IgM responses could not be detected in any of the sera among JRA patients or control children, although they were present in the sera of all 12 (100%) patients with clinical rubella. These data, therefore, fail to suggest serological evidence for the association of RV infection with JRA.

BK virus infections in children with various primary immunodeficiencies and in related healthy household contact persons

Sera from 26 patients with different inherited immunodeficiences were tested for specific antibodies to BK virus (BKV) in the hemagglutination inhibition (HI) and the IgM — immunofluorescence (IF) test. Eighty-one percent exhibited HI-antibodies to BKV, whereas virus-specific IgM antibodies were not found in a single case. Serial serum samples from seven patients did not show significant titer changes during the observation period of up to two years. A comparison was made between the antibody levels of the sera tested and the occurrence of BKV in the corresponding urine samples. Furthermore, in eight patients the seroreactivity of related close household contact persons was investigated. Seven of the 17 healthy relatives investigated had BKV-specific IgM antibodies and most of them revealed high serum antibody levels in the HI-test.

Primary Epstein-Barr virus infection in early childhood.

In a prospective study 43 consecutive children in hospital, aged between 6 months and 7 years and displaying at least one of the clinical signs of infectious mononucleosis (IM), were investigated for Epstein-Barr (EB) virus-specific IgM antibodies by an indirect immunofluorescence test. On this basis EB virus infection was considered confirmed in 8 patients, each of whom had IgM antibodies in the initial serum sample. In one additional patient, IgM antibodies were only detected in a second sample. The IgM antibodies disappeared with 3-11 weeks. Assessment of IgG antibodies had no diagnostic value in the acute phase of IM. Clinically the 3 youngest children, about 1 year of age, were diagnosed as having pneumonia or hepatitis, the 5 other consecutive patients as having IM. Hepatosplenomegaly was fairly frequently associated with IM, while sore throat, lymphadenopathy, and rash were often signs of other diseases. Only the oldest child had heterophil antibodies. Atypical lymphocytes (greater than 10%) were present in 4 of the 9 IM cases and were seen in children with other diseases as well. Our data stress the importance of measuring EB virus-specific IgM antibodies in order to diagnose IM in early childhood.

Enzyme-linked immunosorbent assay (ELISA) for mumps virus antibodies.

An ELISA for the detection of mumps virus-specific IgG and IgM antibodies was developed. Three different antigen preparations were compared. Equally good results were obtained with virus concentrated by ultracentrifugation and with virus that was further purified by sucrose gradient centrifugation. Crude infected allantoic fluid was unsuitable for use an antigen. The variability and reproducibility of the tests were satisfactory. When the ELISA was compared to conventional serological methods, a good correlation of IgG absorbance values with complement fixation (CF) antibody titers was found (r = 0.574), the ELISA being more sensitive in detecting antibodies in acute-phase sera. For the determination of immunity, ELISA IgG values were compared with results obtained in hemagglutination-inhibition (HI) and hemolysis-in-gel (HIG) tests. Again there was a good correlation with both tests (rHI = 0.528, rHIG = 0.667). The ELISA was more sensitive than the HI and HIG test for the detection of low levels of antibodies.

Detection of IgM Antibodies to Cytomegalovirus (CMV) Using an Enzyme-labelled Antigen (ELA)

We have applied a peroxidase enzyme-labelled antigen (ELA) for the detection of IgM antibodies to cytomegalovirus: microtitre plates were coated with anti-IgM immunoglobulin. The IgM fraction of human serum was selectively bound to the precoated plates and the virus-specific IgM antibody was then detected by the enzyme-labelled antigen. A very efficient technique for the labelling of virus antigen is described. The IgM antibody was detected simply and specifically, Rheumatoid factor IgM did not interfere with this test.

Detection of enterovirus specific IgG and IgM antibodies in humans by an indirect solid phase radioimmunoassay.

The development of a solid phase radioimmunoassay which is able to detect virus-specific IgG and IgM antibodies in serum specimens from patients with enterovirus infections is described. Viral antigen partially purified from infected tissue culture fluid was absorbed passively to individual polystyrene microtiter wells. Dilutions of sera were incubated on these antigens and bound anti-viral antibodies were monitored by the addition of 125-iodine labeled anti-human-IgG or anti-human-IgM antibody. Specificity of the assay to detect virus-specific IgM antibody was ensured by highly specific anti-IgM antibody which did not cross react with IgG and 2-mercaptoethanol sensitivity of IgM antibody titers. Changes of IgM antibody titers clearly indicated a current infection by that virus strain which was isoalted as etiological agent. Advantages and restrictions of the introduced radioimmunoassay are discussed.

A new method for the determination of virus specific IgG and IgM antibodies

A Latex test has been developed for the detection of virus specific IgG and IgM antibodies. Virus was incubated with serum and the resulting virus-antibody complex harvested. Latex particles sensitized with anti IgG and anti IgM were added and the quantity and type of bound antibody determined.

Measles virus-specific IgM antibodies and IgM-class rheumatoid factor in serum and cerebrospinal fluid of subacute sclerosing panencephalitis patients

Serum and cerebrospinal fluid (CSF) from 20 subacute sclerosing panencephalitis (SSPE) patients were analyzed with solid-phase radioimmunoassays designed to detect measles virus-specific IgM antibodies and IgM-class rheumatoid factor (RF). Serum and CSF from patients 1 to 16 did not contain measles IgM antibodies or elevated levels of RF. Serum from all and CSF from two of the remaining four patients were apparently measles IgM positive. RF levels were elevated, however, in serum from two and in serum and CSF from another of these four patients. Testing after removal of IgG or RF revealed that patient 17 had both measles IgM and elevated levels of RF in serum, and measles IgM in CSF. Reactivation of measles IgM synthesis was detected in the blood of patient 18. The serum of patient 19 and the serum and CSF of patient 20 contained only elevated levels of RF. The implications of these results are twofold. First, although measles IgM is found in some SSPE patients, it is not found consistently enough to be a marker for the chronic measles virus infection in these patients. Second, the possible interfering presence of RF must be taken carefully into account in any study of IgM antibody persistence.