Evaluation of five commercial enzyme immunoassays for the detection of human cytomegalovirus-specific IgM antibodies in the absence of a commercially available gold standard.

In the recent years the number of commercially available immunoassays for the detection of human cytomegalovirus (HCMV)-specific immunoglobulin M (IgM) antibodies has rapidly increased. The aim of the present study was to evaluate five commercial immunoassays for the serological diagnosis of HCMV-infection. These methods, namely the IMx CMV IgM assay, the AxSYM CMV IgM assay (both Abbott), the Gull CMV IgM, the CMV-IgM-ELA test PCS Medac and the Biotest Anti-HCMV recombinant IgM ELISA, were compared for their diagnostic effectiveness and interference with substances eventually producing cross-reactions with HCMV-IgM (Epstein-Barr-virus (EBV)-IgM, rheumatoid factor (RF)). In addition, repeated measurements on samples from kidney and heart transplant recipients with active HCMV infection were examined to compare the temporal development of the HCMV-IgM measured with the five assay systems. Since there is no commercially available gold standard, it was assumed that the true classification, of whether the patient sample is HCMV-IgM positive or negative, was unknown. Hence sensitivity and specificity were assessed based on a maximum likelihood approach using a "latent class" model. The cross-reactions were quantified by a Bayesian statistical model using prior information for the expected prevalences in the EBV-IgM and rheumatoid factor sample groups. The results of the study demonstrated that there are great differences in sensitivity and specificity as well as in cross-reactions with EBV-IgM and RF between the tested ELISAs.     

An evaluation of five commercially available kits for the diagnosis of rotavirus infection

A total of 172 faecal specimens were examined by Rotazyme II, Wellcozyme Rotavirus, Wellcome Rotavirus Latex, Mercia Rota Screen and Bio Merieux Slidex Rota Kit. Assuming a true positive to be when two or more test systems were in agreement, analysis showed that 19 specimens gave positive results. Rota Screen proved to be the most consistent and reliable of the latex kits giving two false negatives, no false positive reactions, and no nonspecific reactions. Wellcome Rotavirus Latex showed one false negative and four non-specific reactions but 14 false positive reactions were observed. The Slidex Rota Kit proved insensitive on initial testing, detecting only nine of the positive specimens and showing thirteen “non-specific” reactions. On further examination after the filtration of specimens which gave aberrant results and testing with a different batch of reagents non-specific results were reduced to five but only 15 specimens were considered to give positive reactions. Both enzyme immunoassay (EIA) test systems proved to be slightly more sensitive than latex tests each producing positive results in 18 specimens. Rotazyme II, however proved to be less specific than Wellcozyme Rotavirus as four false positive reactions were noted, only one false positive reaction being detected with Wellcozyme Rotavirus. Both EIA methods failed to detect one positive specimen.     

HCMVpp65抗原和IgM抗体检测诊断儿童HCMV活动性感染的比较

目的：比较人巨细胞病毒(HCMV)pp65抗原(以下简称pp65)检测和血清HCMV-IgM抗体(以下简称IgM抗体)检测两种方法对儿童HCMV活动性感染的诊断实用意义。方法：采集临床疑似HCMV活动性感染儿童血标本(共251份)，分离血浆和多形核白细胞，分别用于IgM抗体检测和pp65检测。同时进行荧光定量PCR检测HCMVDNA，与pp65抗原作平行比较。结果：pp65抗原检测的结果与IgM抗体检测的符合率为73.3％。与HCMV DNA检测相比pp65抗原检测法的符合率、特异度和敏感度分别为85．5％，85．2％和86．2％。而且高pp65抗原血症与患者的临床症状密切相关。结论：pp65抗原血症反映该病毒活动状况，可监测HCMV活动性感染。由于儿童患者还存在原发感染的可能性，所以．联合HCMV-IgM的监测来提高临床的诊断率。     

Evaluation of a novel, commercially available mass spectrometry kit for newborn screening including succinylacetone without hydrazine

Newborn screening for tyrosinemia type I (Tyr-I) is mandatory to identify infants at risk before life-threatening symptoms occur. The analysis of tyrosine alone is limited, and might lead to false-negative results. Consequently, the analysis of succinylacetone (SUAC) is needed. Current protocols are time-consuming, and above all, include hazardous reagents such hydrazine. We evaluated a novel, commercial kit to analyze amino acids, acylcarnitines and SUAC with a significantly less harmful hydrazine derivative in a newborn screening laboratory. Dried blood spot specimens from 4683 newborns and samples from known patients with inborn errors of metabolism (IEM) were analyzed by a novel protocol and compared to an in-house screening assay. All samples were derivatized with butanol-HCl after extraction from 1/8-inch DBS punches. For the novel protocol, the residual blood spots were extracted separately for SUAC, converted into hydrazone, combined with amino acids and acylcarnitines, and subsequently analyzed by mass spectrometry using internal isotope-labeled standards. All newborns were successfully tested, and 74 patients with IEMs including three with Tyr-I (SUAC 1.50, 4.80 and 6.49; tyrosine levels 93.10, 172.40 and 317.73, respectively) were detected accurately. The mean SUAC level in non-affected newborns was 0.68 μmol/l (cut-off 1.29 μmol/l). The novel assay was demonstrated to be accurate in the detection of newborns with IEM, robust, and above all, without the risk of the exposure to highly toxic reagents and requirement of additional equipment for toxic fume evacuation.     

Evaluation of four commercially available ELISA assays for the serologic diagnosis of Lyme disease

We evaluated four commercially available ELISAs for detection of antibody to Borrelia burgdorferi with 21 sera from patients with clinically diagnosed Lyme disease and 89 patient control sera. Patient control sera included 28 sera from patients with rheumatoid arthritis (RA), 17 sera from patients with systemic lupus erythematosus (SLE), and 44 sera containing antibodies reported to cross-react in some Lyme disease tests. The ELISAs tested (Cambridge Bioscience, Diamedix, 3M, and Zeus) detect antibodies (IgM and/or IgG) that bind Borrelia burgdorferi antigen attached to microtiter wells. Antibody reactivity in the sera from patients with clinically diagnosed Lyme disease was characterized by using Zeus immunoglobulin class-specific assays (IgM and IgG). Sensitivities in early and late Lyme disease were as follows: Cambridge and Diamedix, 57% and 100%; 3M, 57% and 93%; and Zeus, 71% and 86%. Reactivities within a patient control population were: Cambridge and Diamedix, 3%; 3M, 7%; and Zeus, 10%.     

Evaluation of three commercially available kits for serological diagnosis of dengue haemorrhagic fever

Seventy one acute phase serum samples collected during an epidemic of dengue hemorrhagic fever were tested by immunoblot, a rapid immunochromatographic assay and Dengue Duo ELISA for presence of anti dengue IgM and IgG antibodies. A concordance of 81.7% and 76.1% was seen between the three tests for the detection of anti-dengue IgM antibodies and IgG antibodies respectively. The rapid test takes only five minutes, can be easily carried out in most laboratories and compares well with the ELISA and the immunoblot.     

Evaluation of the human basophil degranulation test using the commercially available Baso-kit as a test of immediate-type hypersensitivity in hay-fever sufferers

The basophil degranulation test (BDT) has failed to gain popular acceptance as a test of immediate-type hypersensitivity since its introduction in clinical medicine in 1961. Recent modification of this technique, however, seemed to have increased its reliability. We have studied a group of twenty-five hay-fever sufferers and a similar age- and sex-matched group of healthy controls to compare the sensitivity of a newly available commercial BDT kit (Baso-kit, Laboratoire des Stallergenes) with specific serum IgE levels and skin-prick tests to meadow-grass pollen. A quantitative correlation was found between the positive BDT (greater than 30% degranulation) and the skin tests, whereas only a qualitative relationship was obtained with specific IgE concentrations. The BDT also identified two subpopulations of responders and non-responders among a third group of hay-fever subjects who had previously been hyposensitized to mixed grass pollen. We conclude that the modified BDT provides a worthwhile addition to the in vitro testing of immediate-type hypersensitivity states.     

Evaluation of four commercially available assays for free thyroxin

We assessed two two-step and two analog assays for measuring free thyroxin (FT4) in serum: Clinical Assays' "GammaCoat Free/Total T4" (CA), Vitek's "KinetiCount Phase II Free T4" (VTK), Diagnostic Products Corporation's "Coat-a-Count Free T4" (DPC) kit (June 1987 version), and Amersham's "Amerlex-M Free T4" (AMX). The VTK assay is automated except for the initial pipetting step. Interassay results correlated well except for samples with abnormal serum albumin concentrations. FT4 values for hypoalbuminemic samples showed a highly significant (P less than 0.0001) correlation with serum albumin concentration in the DPC and AMX assays. The relationships are described by the equations y = 0.382albumin (g/L) + 0.81 pmol/L and y = 0.450albumin (g/L) - 3.20 pmol/L, respectively. When we used an equation derived from the Law of Mass Action to adjust FT4 values to values expected at an ideal albumin concentration, the observed correlation of albumin and FT4 was abolished completely in the DPC assay, and partly so in the AMX assay. The precision of CA was comparable with that of the analog assays; the CV for the VTK assay was approximately twice that for the other three assays.     

Adaptation of a commercially available indirect fluorescent antibody slide test for measuring measles-specific immunoglobulins

A commercially available indirect fluorescent antibody (IFA) test was evaluated for the determination of measles-specific immunoglobulins G and M (MIgG, MIgM). The IFA test detected fourfold rises of MIgG in 34 of 35 (97%) cases of measles confirmed by complement fixation or hemagglutination inhibition. In determining immune status, MIgG-IFA correlated with hemagglutination inhibition in 22 of 23 (96%) cases. The IFA test detected MIgM in only 11 of 34 acute-phase sera collected within 5 days of the reported onset of rash and in the convalescent specimens of another 13 of the 35 specimens. The IFA test is an effective method for the conventional diagnosis of measles and for determining immune status. This IFA test has a limited role as a rapid diagnostic test for measles when used to detect measles-specific MIgM in acute-phase sera obtained from patients with suspected measles.     

Evaluation of a commercially available ELISA assay for detection of Giardia lamblia in fecal specimens

A total of 417 fecal samples preserved in 10% buffered formalin and PVA were submitted to a commercial microbiology laboratory only for the detection of Giardia lamblia. Results from fecal specimens collected from 411 patients with gastrointestinal symptoms were compared using the following methods: (a) standard Ova & Parasite (O&P) concentration; (b) Alexon's ProspecT/Giardia enzyme-linked immunosorbent assay (ELISA) test, and (c) Meridian's Direct Fluorescent Antibody (DFA) Stain. In the 29 specimens in which G. lamblia was detected, 10 were O&P, DFA and ELISA positive, 17 were only ELISA positive and two were only Ova & Parasite and Direct Fluorescent Antibody positive. Of the 29 positive specimens, 22 were confirmed as true positives. The ELISA sensitivity was 91% and the specificity was 98%. The expense associated with these methods to detect the presence of Giardia is $11.00, $8.95, and $12.80, respectively. In symptomatic patients, the ProspecT/Giardia ELISA is a cost-effective, rapid, and sensitive method for detecting the presence of G. lamblia in fecal specimens.     

4种商品化支原体液体培养试剂盒的质量评价

目的比较4种商品化支原体液体培养鉴定检测试剂盒,评价不同厂家支原体试剂的应用效果。方法在相同条件下,选择4种支原体液体培养鉴定试剂和固体培养法对90例保存的临床标本分别进行检测。结果以固体培养法结果为“金标准”,按试剂说明要求,4种支原体液体培养试剂24小时对解脲支原体(Uu)的敏感性和特异性分别为:进口试剂IST为96%和100%、国产试剂1为44%和100%、国产试剂2为68%和100%、国产试剂3为18%和100%;48小时对人型支原体(Mh)的敏感性和特异性分别为:进口试剂IST为95%和100%、国产试剂1为65%和100%、国产试剂2为2.5%和100%和国产试剂3为0%和100%。固体培养结果与进口IST支原体液体培养试剂结果比较差异无统计学意义(P>0.05),但是与国产的3种试剂结果比较差异均有统计学意义(P<0.01)。结论进口IST支原体液体试剂培养鉴定结果与固体培养结果符合率最高,其他3种试剂与固体培养结果符合率较差,结果提示临床检测时,选用支原体液体培养鉴定试剂应慎重。     

Evaluation of two commercially available photometers for blood donor haemoglobin measurement

Two commercially available portable haemoglobin measuring devices (Delphi Haemoglobin Meter and HemoCue) were evaluated both within the laboratory and under routine operating conditions in blood donation collection centres. Both meters demonstrated satisfactory accuracy when compared to the reference cyanmethaemoglobin (HiCN) method. Within-run and between-run precision of both meters was satisfactory. There was no significant difference evident between capillary and venous haemoglobin results using either meter. In the main blood collection centre, using Delphi meters, 63.1% of results returned from an external proficiency testing program were acceptable (± 2SD of Quality Control Laboratory result). In satellite centres using Delphi meters 36.5% were acceptable while 60.0% of HemoCue results from satellite centres were acceptable. Assay time was almost twice as long using the HemoCue. The study confirmed the necessity to evaluate equipment and operators under both controlled and routine operating conditions. External proficiency testing programs are recommended to monitor operator performance on a regular basis.     

Evaluation of a commercially available serologic assay for antibodies against tuberculosis-associated glycolipid antigen.

A commercially available enzyme immunoassay developed to detect antibodies to a tuberculosis-associated glycolipid antigen was evaluated for serologic diagnosis of tuberculosis. This was a multicenter study comparing the assay with other methods in 78 patients with active pulmonary tuberculosis and in 54 controls with non-tuberculous lung diseases. Sensitivities were highest for sputum culture (91.0%), followed by immunoassay (79.5%), nucleic acid amplification (77.3%), and finally acid-fast staining of sputum smear (60.3%). Immunoassay and amplification, both rapid methods, had similarly high sensitivity in smear-positive subjects (89.4 and 88.9%, respectively); in smear-negative subjects these two techniques showed low sensitivity (64.5 and 60.0%, respectively). Concordance between the two methods was relatively low (72.0%). With regard to specificity, seven out of ten patients with old tuberculosis had positive result by immunoassay (30% specificity). In the control group, 10 out of 54 patients had positive immunoassay result (72.2% specificity), with notably limited specificity in the elderly. The tuberculous glycolipid assay is a rapid method sufficiently sensitive for detection of tuberculosis infection, even in smear-negative patients.     

Prenatal diagnosis of cytomegalovirus infection

Prognostic value of markers of cytomegalovirus infection (CMV) in pregnant women for the neonatal status was assessed. Detection of such markers as antiCMV IgM and CMV DNA in cervical secretion by DNA dot-spot hybridization in women with a complicated course of pregnancy indicates a 5.7% risk of delivery of children with stable symptoms. Studies of antibodies to pre-early proteins (IE CMV) showed that antiCMV IgG to IE are more incident in pregnant women than antiCMV IgM; moreover, antiCMV IgG to IE but not antiCMV IgM are detected in umbilical blood. The results of detection of antiCMV IgG and IgM to IE correlated with the clinical characteristics of newborns.     

Comparison of the detection of Helicobacter pylori infection by commercially available serological testing kits and the 13C-urea breath test

BackgroundSerum Helicobacter pylori (H. pylori) antibody kits (LZ and LIA) using the latex agglutination immunoassay method are commercially available, but few studies have been performed to determine their diagnostic accuracy or to compare their results with those of enzyme-linked immunosorbent assay (ELISA) kits (EP and EIA).MethodsSera were obtained from 213 hospital outpatients with dyspeptic symptoms. The serological results were compared with the result of the ¹³C-urea breath test (UBT) which seems to be reliable.ResultsOf the 213 subjects, 154 were diagnosed as positive for H. pylori infection according to the UBT. The sensitivities and specificities of these tests were 97.4% and 76.3%, 98.1% and 78.0%, 99.4% and 74.6%, and 98.1% and 71.2% for the EP, LZ, EIA and LIA tests, respectively. When the 13 subjects whose seropositive results of the four kits were completely opposite to the negative results of the UBT were excluded, the specificities of evaluated kits were all higher than 90%. The concordance rate between the EP and EIA tests was 98.1% (Spearman's rank correlation coefficient = 0.83) and that between the LZ and LIA tests was 97.1% (correlation coefficient = 0.91). The LZ gave higher antibody titer value than EP (p < 0.0001, Z = 9.82; Wilcoxon signed-rank test), and EIA gave higher value than LIA (p < 0.0001, Z = 6.43; Wilcoxon signed-rank test).ConclusionsThe latex immunoassay method provided the same reliability to ELISA in terms of the diagnostic accuracy for current H. pylori infection, although we should take into account the titer value differences by each test method in practical use.