Immunoglobulin Classes of Antibody for Human T-Lymphotropic Virus Type-I (HTLV-I) in Healthy Donors and HTLV-I-Associated Disorders

Healthy blood donors, patients with adult T-cell leukemia (ATL) and HTLV-I-associated myelopathy (HAM) and recipients of unscreened blood (SR) who had seroconverted and were followed-up for more than 2 years were examined for HTLV-I antibodies of immunoglobulin G (IgG) and M(IgM) classes. The overall infection rate in donors was 4.9%, as determined by screening with a particle agglutination method (PA). The rate increased with increasing age. Positive sera with a low titer in the PA test (1/16, 1/32 and 1/64) contained IgM antibodies in 32.5% (titer 1/16) to 36.1% (titer 1/64) of the cases, but IgG antibodies were detected in only 5.6% of the sera with a titer of 1/16 and in 36.1% of the sera with a titer of 1/64. Conversely, in high titer sera (1/128 or higher) IgG antibodies were almost always detectable (99.0%) and IgM antibodies less frequently (25.5%). Sera from acute, chronic and pre-ATL, HAM and SR patients contained IgG antibodies in high titer in all cases. The incidence of IgM antibodies was 7.7, 30.0, 53.3, 72.3, and 77.8%, respectively. IgM antibodies were demonstrated repeatedly in some cases who were followed up for a year. Only IgM antibodies from HAM patients occurred in high titers and had strong reactivity to the p24 antigens of HTLV-I in Western blot testing. It is concluded that it is important to detect IgM antibodies not only in primary infections but also in persistent infections of HTLV-I.     

Immunoglobulin M antibodies are not specific for serodiagnosis of human toxocariasis

Serodiagnosis of human toxocariasis is established by detecting serum anti‐Toxocara IgG antibodies, but there is little knowledge regarding the reactivity of human IgM antibodies against the Toxocara antigens. In this study, we have evaluated the reactivity of IgM antibodies in sera from patients with toxocariasis, patients with other helminth infections, and healthy individuals, against Toxocara larval excretory‐secretory (TES) antigens by enzyme‐linked immunosorbent assay (ELISA) and Western blot (WB). Anti‐Toxocara IgM were detected in 91.4% of sera from patients with toxocariasis, 76% of sera from patients with other helminth infections, and 45.3% of sera from healthy individuals when ELISA was used. Likewise, IgM antibodies were detected in 94.8% of sera from patients with toxocariasis, 65.3% of sera from patients with other helminth infections, and 41% of sera from healthy individuals when WB was used. This reactivity exhibited only a slight decrease when the TES antigens were deglycosylated, showing that not only glycosidic epitopes, but also peptide epitopes are involved in the recognition and binding of IgM antibodies during the immune response against the parasite. The results shown that IgM antibodies are not specific for serodiagnosis of human toxocariasis.     

HIV-1 infection in HIV-1 enzyme-linked immunoassay seronegative patients in Kinshasa, Zaire

Serum samples of 62 African patients who had clinical manifestations of HIV-1 infection but were seronegative for HIV-1 by ELISA (Organon) were subsequently further tested by another HIV-1 ELISA test (Wellcozyme), HIV-1 IgG Western blot, HIV-1 antigen detection and HIV-2 ELISA. Patients' lymphocytes were cultured for HIV-1 and 2. Because of limited quantities of serum available all tests were not performed on all samples. Seven (26%) of 27 sera of patients meeting the WHO clinical case definition of AIDS were Western-blot-positive. In contrast, of 35 patients' sera with possible HIV related disease, only one (3%) was Western blot positive (P = 0.02) and none of 75 sera from HIV-1 ELISA (Organon) seronegative blood donors (P less than 0.01) were Western blot positive. Of 30 HIV-1 ELISA (Organon) seronegative patients tested with the HIV-1 ELISA Wellcozyme assay only one was seropositive (this patient's serum was also Western blot positive). Of 17 HIV-1 ELISA (Organon) seronegative patients tested, HIV-1 antigen was found in 1 case (6%) (this patient's serum was Western blot negative). None of the 34 patients tested by HIV-2 serology was HIV-2 seropositive. HIV-1 was isolated by culture in 3 (21%) of 14 HIV-1 ELISA seronegative patients (sera of the 3 patients were Western blot negative). In total, 12 (19%) of 62 HIV-1 ELISA (Organon) seronegative patients were found to be positive for HIV, either by Western blot HIV antigen testing or viral culture.(ABSTRACT TRUNCATED AT 250 WORDS)     

Frequency and specificity of antibodies that crossreact with Borrelia burgdorferi antigens.

The frequency and specificity of antibodies that bind antigens of Borrelia burgdorferi in sera from 200 individuals with no evidence of past or current Lyme disease was determined. Sera were tested for both IgG and IgM antibodies to B. burgdorferi by Western blotting. The non-Lyme serum group included specimens from healthy adults and children in addition to specimens from patients with viral infection and rheumatic diseases. Crossreactive IgG antibodies occurred more frequently than IgM antibodies. The most frequently bound antigens corresponded to 41 kDa and 60 kDa Borrelial components. Of 200 specimens tested, 100 had antibodies that bound at least 1 antigen. Binding to multiple antigens occurred at much lower frequency. Our results indicate that determination of maximum crossreactivity of non-Lyme sera can be used to establish minimum criteria for determining a positive Western blot result for Lyme disease.     

Simultaneous detection of IgM antibodies against the hepatitis A virus and the viral capsid antigen of Epstein-Barr virus in acute hepatitis

The simultaneous detection of IgM antibodies to hepatitis A virus (anti-HAV IgM) and IgM antibodies to viral capsid antigen (anti-VCA IgM) of Epstein-Barr virus (EBV) in patients with acute viral hepatitis has led us to systematically study serological markers of EBV in patients with anti-HAV IgM positive acute hepatitis and to test for anti-HAV IgM in sera of patients with acute hepatitis associated with serological evidence of current primary EBV infection. All patients studied were HBsAg negative and were not drug-addicts, nor homosexuals. In 15 consecutive patients with anti-HAV IgM positive acute hepatitis, anti-HAV IgM and anti-VCA EBV IgM antibodies were simultaneously detected in 9 cases. Of these 9 patients, antibodies to nuclear antigen were positive in 8 cases, antibodies to early antigen were positive in 7 cases and rheumatoid factor was positive in 4 cases. In 5 consecutive patients with acute hepatitis associated with serological evidence of current primary EBV infection, anti-HAV IgM was not detected. Simultaneous presence of anti-VCA EBV IgM, early antigen IgG antibodies and nuclear antigen antibodies in 7 patients with acute hepatitis associated with anti-HAV IgM suggests reactivation of EBV or reactivation of clones secreting antibodies anti-EBV in HAV infections. Furthermore, these results show that anti-VCA IgM only cannot be considered to be a specific marker of early EBV infection in patients with acute hepatitis.     

伯氏疏螺旋体与苍白密螺旋体、钩端螺旋体的免疫交叉反应抗原研究

目的研究莱姆病螺旋体与梅毒、钩端螺旋体较常出现的交叉反应抗原，明确产生交叉反应的蛋白抗原成分，为精确蛋白免疫印迹法检测莱姆病的阳性判断标准提供参考依据。方法以中国莱姆病螺旋体伽氏疏螺旋体基因型代表菌株PD91作抗原，用蛋白免疫印迹法对梅毒病人和钩端螺旋体病人的血清进行抗体(IgG和IgM)检测。结果共检测梅毒病人血清196份和钩端螺旋体病人血清68份。伯氏疏螺旋体与苍白密螺旋体抗原在75kDa，60kDa，43kDa，41kDa处有交叉，交叉反应阳性率分别为IgG：8．2％、20．4％、9．2％、17．3％，IgM：7．7％、10．2％、8．7％、9．7％；伯氏疏螺旋体与钩端螺旋体抗原的交叉发生在75kDa，60kDa，41kDa，交叉反应阳性率分别为IgG：1．5％、1．5％和13．2％，IgM：8．8％、2．9％、17．6％。结论莱姆病螺旋体蛋白抗原中75kDa，60kDa，43kDa和41kDa蛋白成分与梅毒和钩端螺旋体存在交叉反应。     

Human parvovirus B19 infection in patients with systemic lupus erythematosus

OBJECTIVE: The clinical significance of the presence of B19 DNA in patients with SLE was studied. METHODS: Sera from 72 patients with systemic lupus erythematosus (SLE), 23 patients with rheumatoid arthritis (RA), 18 patients with Sj?gren's syndrome (SS), eight patients with Raynaud's phenomenon (RP), five patients with primary biliary cirrhosis (PBC), five patients with polymyositis (PM), four patients with erythema infectiosum (EI) and 22 normal controls were examined for parvovirus B19 (B19) infection by serological assays, nested PCR and Southern blotting. RESULTS: Parvovirus B19 DNA was detected in 17 of 72 patients with SLE and in three of four patients with EI, but not in patients with other systemic rheumatic diseases. Of the 17 patients with B19 DNA, only one had IgG anti-B19 antibody and two had IgM anti-B19 antibodies, whereas IgG and IgM anti-B19 antibodies were detected in 27 (49.1%) and 21 (38.2%) of 55 SLE patients without B19 DNA respectively. All sera from the patients with EI contained both IgG and IgM anti-B19 antibodies. B19 DNA was found more commonly in sera from SLE patients without anti-B19 antibodies than in those with anti-B19 antibodies (P<0.05). CONCLUSIONS: B19 infection in patients with SLE may be due to lack of anti-B19 antibodies because of either the immunocompromised nature of the host or the use of immunosuppressive drugs. There was a higher prevalence of hypocomplementaemia and RP in patients with parvovirus B19 viraemia than in those without parvovirus B19 viraemia.     

Trypanosoma cruzi: a carbohydrate epitope defined by a monoclonal antibody as a possible marker of the acute phase of human Chagas' disease.

An IgM monoclonal antibody (MAb) against a carbohydrate epitope present in Trypanosoma cruzi trypomastigote excretory-secretory antigens and expressed by different developmental stages of the parasite (epimastigote, trypomastigote and intracellular amastigote) was linked to a solid phase matrix and used as an antigen-capture antibody. Human serum complexes containing the epitope were then detected by using specific secondary antibodies against human immunoglobulin isotypes. Results of detection of IgM, IgG, and IgA serum complexes (SC) containing a T. cruzi polypeptide epitope showed that SC could be detected in 69% of the 13 Chagasic acute phase sera studied with IgG, in 84% with IgM, and in 75% with IgA. Only 16% (IgG-SC), 8% (IgM-SC), and 10% (IgA-SC) of chronic sera from 50 patients were positive. No patients with toxoplasmosis or rheumatoid factor were positive. Of the 11 leishmaniasis sera studied, four had IgG-SC, two had IgA-SC, and five had IgM-SC. Of the eight Yanomamo Indians infected by Onchocerca volvulus, three were found to have IgG-SC, two had IgM-SC, and two had IgA-SC. Thirteen sera from healthy individuals living in an endemic area were also studied. One subject had IgG IgM and IgA-SC. The results presented in this study show for the first time, the specific detection of IgM, IgG, and IgA immune complexes using a MAb against T. cruzi. The presence of the epitope in association with IgM antibodies in sera from patients with the acute phase of the disease suggests that this antigen(s) carrying the epitope that reacts with the MAb could be a marker(s) of active infection. In addition, the specificity of the serum complex capture assay allowed the detection of Chagas' disease in two different endemic areas (Argentina and Venezuela).     

Clinical evaluation of commercial serological test for Bartonella infection

We evaluated the usefulness of a serological diagnostic kit (Bartonella IFA IgG, IgM; MRL Diagnostics) for Bartonella henselae infection. Of the 110 healthy individuals, 107 (97.3%) were with titers being less than 1:64 for IgG antibody to B. henselae, 2 were with titers being 1:64 and 1 with 1:128, IgM antibody to B. henselae was negative in all individuals. Serological diagnosis of cat scratch disease (CSD) using indirect fluorescence antibody (IFA) methods (in-house and diagnostic kit) was made in either elevated titers of IgM (> or = 1:20) or IgG (> or = 1:256) antibodies, or a four-fold rise in IgG titer between acute and convalescent sera. Of the 18 individuals with serological diagnosis of CSD by in-house IFA method in 26 CSD clinical diagnosed patients, 15 (83%) were compatible with the results of the diagnostic kit, whereas 3 (17%) were not compatible. Of the 8 without serological diagnosis, 1 (13%) was serologically diagnosed as CSD, and the others were negative. Overall, the serological diagnosis was made in 16 of 26 (62%). The specificity and sensitivity of this kit were 100% and 62%, respectively. The cross-reaction between B. henselae and Bartonella quintana was observed in sera from controls and patients. Our results show that the diagnostic kit as well as in-house method is an useful tool for the serological diagnosis of cat scratch disease.     

肺孢子虫病实验与临床研究 Ⅱ．免疫印迹试验检测人血清肺孢子虫抗体

采用ＳＤＳ－ＰＡＧＥ分析鼠源卡氏肺孢子虫包囊可溶性抗原呈现２０条以上多肽区带，主带分子量分别为＞１００、８５－９４、６７、５２和３４ｋＤａ。ＥＩＴＢ检测表明重庆地区健康人血清中存在识别１１０、１０５、８５、６７、５２和４６ｋＤａ的ＩｇＧ型抗体，抗体阳性率为５６．７％（５９／１０４）。其中８５ｋＤａ抗原能与抗人源肺孢子虫单克隆抗体２Ｇ２起反应。健康人血清抗８５ｋＤａ抗原的抗体阳性率为３７．５％（３９／１０４），但ＩｇＭ型抗体均为阴性。另外检测７例确诊的卡氏肺孢子虫病患者，４例血清ＩｇＧ型抗体阳性，其中３例抗８５ｋＤａＩｇＧ型抗体阳性。     

Detection of cross-reacting idiotypes in sera of lymphoma patients by inverse monoclonal radioimmunoassay

An anti-idiotypic IgG1 kappa murine monoclonal antibody (MoAb) Y7 against purified monoclonal IgM lambda 1, derived from a patient with Waldenstr?m's macroglobulinemia, has been generated. This antibody cross-reacted with the tumor-derived idiotypes of patients with B cell non Hodgkin's lymphoma as measured by competitive inverse solid radioimmunoassay using unpurified serum samples. Our results with the inhibition curves of 10 sera of normal donors and 60 sera of lymphoma patients indicate that 21 lymphoma patients revealed cross-reactivities greater than 7%, the mean value observed in normal donors. Of these, 5 sera cross-reacted strongly, in the range of 43-163%, revealing a frequency of positive cross-reactivity for MoAb Y7 of 1/12 sera of lymphoma patients. The generation of a panel of anti-idiotypic antibodies which cross-react with different tumor-derived Ig in serum may be valuable for monitoring the disease in a high proportion of NHL patients.     

Seroprevalence of West Nile virus and tick-borne encephalitis virus in southeastern Turkey: first evidence for tick-borne encephalitis virus infections

West Nile virus (WNV) and tick-borne encephalitis virus (TBEV) are among the medically important Flaviviruses that cause significant morbidity and mortality in humans. In this study, seroprevalence of WNV and TBEV in sera from two state medical hospitals from the southeastern part of Turkey was investigated. One hundred eighty-one serum samples were evaluated for WNV immunoglobulin G (IgG) by an indirect immunofluorescence test (IIFT) and for IgG antibodies against TBEV by a commercial enzyme-linked immunosorbent assay (ELISA) kit with enhanced sensitivity and specificity. Sera positive for WNV IgG were further analyzed by plaque reduction neutralization assay (PRNA). TBEV IgM was also investigated by ELISA in all seroreactive samples. Of 181 sera, 29 (16%) were positive for WNV IgG by IIFT and 17 of 179 (9.5%) were confirmed by PRNA. Nineteen of 181 (10.5%) sera were detected to have TBEV IgG. Mean titer of TBEV IgG was 43.0 RU/mL (median, 33.9 RU/mL; cutoff: 20 RU/mL). Four samples with WNV IgG antibodies were also positive for TBEV IgG antibodies. TBEV IgM was detected in 9 of 39 (23%) of all seroreactive sera, where IgM positivity were accompanied by IgG for 6 samples. These results suggest the presence of possible human WNV and TBEV infections in southeastern Turkey where vector activity have previously been detected.     

Isotypic analysis, antigen specificity, and inhibitory function of maternally transmitted Plasmodium falciparum-specific antibodies in Gabonese newborns.

An analysis of Plasmodium falciparum-specific antibodies was performed in pairs of maternal and cord sera from Gabon, a region endemic for malaria. All paired sera (n = 59) had P. falciparum-specific antibodies. Immunofluorescence assays detected parasite-specific IgG1, IgG2, and IgG3 in 100% of the tested pairs (n = 26) and IgG4 in 42% of them. The titers of specific IgG2 and IgG3 were significantly lower in cord than in maternal sera. All maternal sera had specific IgM. Of the seven P. falciparum-IgM positive cord sera, six were associated with malaria-related histological placental changes (MRHPC). In addition, higher titers of specific IgG1 in maternal and cord sera and of specific IgG3 in cord sera were associated with MRHPC. Similar P. falciparum antigens were recognized by cord and corresponding maternal sera in radioimmunoprecipitation and Western blot assays (n = 40). Sixteen of 20 cord sera and 15 of 20 paired maternal sera significantly inhibited in vitro parasite growth. The extent of inhibition did not correlate with the titer of specific antibodies. These data confirm the very effective placental transfer of anti-malarial antibodies. The presence of IgM in some cord sera raise the question of intrauterine sensitization to malaria antigens.     

Significance and specificity of antibodies to neutrophils detected by western blotting for the serological diagnosis of primary sclerosing cholangitis.

Antibodies against neutrophils have been detected in sera from patients with primary sclerosing cholangitis and inflammatory bowel diseases either by immunofluorescence or by enzyme-linked immunosorbent assay. To assess primary sclerosing cholangitis-specific antibodies, we examined sera from 30 patients with clinically and morphologically well-established primary sclerosing cholangitis by Western blotting against neutrophils and compared these results with those obtained by testing sera from patients with inflammatory bowel diseases. By Western blot using sonified neutrophils, 24 (80%) of 30 primary sclerosing cholangitis sera were positive. Five antigenic determinants at 95, 60, 55, 40 and 30 kD were visualized. Twenty-eight of the primary sclerosing cholangitis sera also showed the characteristic perinuclear fluorescence pattern by immunofluorescence on neutrophils. Thus a serological diagnosis of primary sclerosing cholangitis could be made in 80% of patients based on these two methods. In contrast, only 9% of 23 patients with ulcerative colitis and 10% of 60 patients with Crohn's disease were positive by Western blot, and these patients also showed positive perinuclear fluorescence pattern by immunofluorescence, suggesting an overlap between inflammatory bowel diseases and primary sclerosing cholangitis. Although some patients with classical primary biliary cirrhosis and autoimmune chronic active hepatitis had antibodies against primary sclerosing cholangitis epitopes, none of the patients with obstructive bile duct disorders, collagen diseases, Wegener's granulomatosis or other hepatic and nonhepatic disorders were positive by Western blot, indicating the specificity of these five primary sclerosing cholangitis-related neutrophilic epitopes.     

Persistence of immunoglobulin M or immunoglobulin G antibody responses to Borrelia burgdorferi 10-20 years after active Lyme disease.

The interpretation of serological results for patients who had Lyme disease many years ago is not well defined. We studied the serological status of 79 patients who had had Lyme disease 10-20 years ago and did not currently have signs or symptoms of active Lyme disease. Of the 40 patients who had had early Lyme disease alone, 4 (10%) currently had IgM responses to Borrelia burgdorferi, and 10 (25%) still had IgG reactivity to the spirochete, as determined by a 2-test approach (enzyme-linked immunosorbent assay and Western blot). Of the 39 patients who had had Lyme arthritis, 6 (15%) currently had IgM responses and 24 (62%) still had IgG reactivity to the spirochete. IgM or IgG antibody responses to B. burgdorferi may persist for 10-20 years, but these responses are not indicative of active infection.     

Use of indirect micromodification of immunoenzyme technique for serologic detection of Ixodes tick-borne borreliosis (comparative analysis)]

Three hundred and seventy one sera sampled in different period since the onset of the disease from 214 patients with Ixodes tick-borne borreliosdes (ITBB) caused by Borrelia afzelii and B. garinii in the Perm Region (Russia) and 299 sera from 222 patients with other diseases were simultaneously tested for IgM and IgG antibodies by using immunofluorescence assay (IFA) with standard antigen from strain Ip-21 B. afzelli by indirect micromodification of enzyme immunoassay (IMELISA) on slides and routine enzyme immunoassay (ELISA) with standard commercial test systems (Medipan Diagnostica GmbH, Germany). It is concluded that IMELISA is a technically easy and accessible test, which is close to IFA in its specificity and sensitivity, and can be widely used for serological verification of ITBB.     

Antigenic types of Orientia tsutsugamushi in Malaysia

The seroprevalence of various Orientia tsutsugamushi (OT) strains among Malaysian patients with suspected scrub typhus infections was determined using an indirect immunoperoxidase (IIP) assay. IgG against a single OT strain were detected in six sera (3 Karp, 1 Gilliam and 2 TC586), whereas IgM antibodies against a single OT strain (Gilliam) were noted in 3 sera (Gilliam). IgG reactive to all OT strains were present in 33 (47.1%) of the 70 sera and IgM reactive to all OT strains were present in 22 (78.6%) of the 28 sera. The fact that most sera were reactive to multiple OT strains suggests that group-specific antigens are involved in scrub typhus infections, whereas very few were due to strain-specific epitopes present on these strains. Peak IgG and IgM titers were noted more frequently against Gilliam, Karp, and TA763 strains: this suggests that these strains may be the commonest infecting strains among Malaysian patients. Two predominant OT polypeptides consistently reacted with patients' sera were the 70 kDa and 56 kDa proteins.     

Serological cross-reaction among Bartonella henselae, Chlamydia pneumoniae and Coxiella burnetii by indirect fluorescence antibody method]

We studied the serological cross-reactions among Bartonella henselae, Chlamydia pneumoniae and Coxiella burnetii by indirect fluorescence antibody (IFA) method, using sera from 8 patients with cat scratch disease (CSD), 13 patients with C. pneumoniae infection and 12 patients with acute Q fever. B. henselae IgG antibody was negative in 13 patients with C. pneumoniae infection, and was positive in 3 (titers being 1:64) of 12 patients with Q fever, whereas B. henselae IgM antibody was negative in all the patients with C. pneumoniae infection or Q fever. C. burnetii IgG antibody was removed by absorption of these 3 sera with C. burnetii antigens, whereas B. henselae IgG antibody did not change. C. pneumoniae IgG antibody was positive in 3 (titers being 1:125 in two, 1:32 in one) of 8 patients with CSD. Both C. pneumoniae and B. henselae IgG antibody titers were significantly reduced by absorption of these 3 sera with B. henselae antigens. C. burnetii IgG or IgM antibodies were negative in all patients with CSD. In conclusion, no serological cross-reaction between B. henselae and C. burnetii was observed. On the other hand. B. henselae IgG antibody cross-reacted to C. pneumoniae antigens, whereas C. pneumoniae IgG antibody did not cross-react to B. henselae antigens. Our findings suggest that determination of B. henselae IgG or IgM antibodies were not influenced by C. pneumoniae and C. burnetii antigens.     

IgM-class antibodies to TT virus (TTV) in patients with acute TTV infection

TT virus (TTV) is a human circovirus with a single-stranded, circular DNA genome of 3.8 kb. A method was developed to detect IgM antibodies to TTV as a serological marker for the diagnosis of acute TTV infection. IgM antibodies in test sera were captured by a monoclonal antibody against IgM/μ on a solid support followed by binding of IgM with TTV derived from fecal extract of a TTV carrier. The presence of IgM-specific TTV particles was determined by polymerase chain reaction (PCR) using nucleic acids extracted from the solid support. Anti-TTV IgM was detected in sera from two patients with non-A to G post-transfusion hepatitis, who were positive for TTV DNA during weeks 10–21 and 12–17, respectively, following transfusion. The anti-TTV IgM was detectable after alanine transaminase levels were elevated and TTV DNA was detectable in the patients. The duration of the anti-TTV IgM was short-lived compared with anti-TTV IgG. Anti-TTV IgM was not detected in sera from any of 36 healthy individuals, with no detectable anti-TTV IgG or TTV DNA in their serum. These results indicate that anti-TTV IgM antibodies would be a useful marker to detect acute TTV infection.