Method for quantifying free hemoglobin, distinct from the hemoglobin-haptoglobin complex, in human serum.

The measurement of free hemoglobin (free Hb) in blood is crucial for assessing the risk of organ damage in patients with hemolytic diseases. However, the colorimetric method, commonly used in clinical practice, does not distinguish between free Hb and the hemoglobin-haptoglobin complex (Hb-Hp) in the blood, instead reflecting the total Hb level. Although size-exclusion high-performance liquid chromatography (SEC-HPLC) can specifically measure free Hb, its clinical use is limited by long assay times. Here, we developed a novel assay method for the rapid quantification of free Hb in serum, distinguishing it from Hb-Hp, using a latex agglutination immunoturbidimetric assay (LATIA). This method could be used to measure free Hb in sera in the range of 1-100 µg/mL in approximately 15 min using an automatic biochemistry analyzer. Using Hb-spiked serum samples from healthy adults, there was a high correlation with Hb levels determined using the newly developed method and SEC-HPLC, indicating a high specificity for free Hb. This novel assay can be used to monitor levels of free Hb in patients with various hemolytic diseases and to design therapeutic strategies based on measured values. However, further studies are required to assess its clinical performance.

Serum Helicobacter pylori antibody reactivity in seven Asian countries using an automated latex aggregation turbidity assay.

BACKGROUND AND AIM: To determine the application range of diagnostic kits utilizing anti-Helicobacter pylori antibody, we tested a newly developed latex aggregation turbidity assay (latex) and a conventional enzyme-linked immunosorbent assay (E-plate), both containing Japanese H. pylori protein lysates as antigens, using sera from seven Asian countries. METHODS: Serum samples (1797) were obtained, and standard H. pylori infection status and atrophy status were determined by culture and histology (immunohistochemistry) using gastric biopsy samples from the same individuals. The two tests (enzyme-linked immunosorbent assay and latex) were applied, and receiver operating characteristics analysis was performed. RESULTS: Area under the curve (AUC) from the receiver operating characteristic of E-plate and latex curves were almost the same and the highest in Vietnam. The latex AUC was slightly lower than the E-plate AUC in other countries, and the difference became statistically significant in Myanmar and then Bangladesh as the lowest. To consider past infection cases, atrophy was additionally evaluated. Most of the AUCs decreased using this atrophy-evaluated status; however, the difference between the two kits was not significant in each country, but the latex AUC was better using all samples. Practical cut-off values were 3.0 U/mL in the E-test and 3.5 U/mL in the latex test, to avoid missing gastric cancer patients to the greatest extent possible. CONCLUSIONS: The kits were applicable in all countries, but new kits using regional H. pylori strains are recommended for Myanmar and Bangladesh. Use of a cut-off value lower than the best cut-off value is essential for screening gastric cancer patients.

Evaluation of the diagnostic potential of recombinant leptospiral OMP A-like protein (Loa22) and transmembrane (OmpL37) protein in latex agglutination test for serodiagnosis of leptospirosis in animals.

Leptospirosis is a re-emerging zoonotic disease of animals and humans caused by pathogenic Leptospira, which has major public health concerns. The study is aimed to express the recombinant outer membrane protein (OMP) A-like protein (rLoa22) and transmembrane (rOmpL37) protein of Leptospira interrogans serovar Hardjo in the Escherichia coli and their evaluation as a diagnostic antigen in the latex agglutination test (LAT) to detect anti-leptospiral antibodies in the sera of animals. The Loa22 and OmpL37 genes lacking signal peptide coding sequences were individually amplified (522 and 963 bp), by polymerase chain reaction, and directionally cloned into a pETite N-His Kan vector for expression. The expressed purified proteins were characterized by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and immunoblot, which confirmed leptospiral specific reactive protein with a molecular weight of ~19 and 36 kDa, respectively. The sensitized latex beads coated with these OM proteins separately were evaluated in LAT using cattle sera of microscopic agglutination test (MAT) confirmed positive (n = 53) and negative (n = 52) cases of leptospirosis. The rLoa22 LAT and rOmpL37 LAT revealed the relative diagnostic sensitivity of 94·34 and 96·23%, diagnostic specificity of 92·31 and 96·15% and accuracy of 93·33 and 96·19%, with the excellent agreement of Cohen's kappa value of 0·87 and 0·92, respectively. After extensive evaluation, this rapid recombinant protein-based field diagnostic test can be applied as a screening test for the detection of anti-leptospiral antibodies in the sera of animals in the field conditions.

Contrasting Results from Two Commercial Kits Testing for the Presence of Clostridium perfringens Enterotoxin in Feces from Norovirus-Infected Human Patients.

BACKGROUND: Detection of Clostridium perfringens enterotoxin (CPE) is critical for disease surveillance; however, commercial testing kits produce contrasting results. METHODS: We examined the cause of the differing results from a reversed passive latex agglutination (RPLA) assay (PET-RPLA Toxin Detection Kit) and an enzyme-linked immunosorbent assay (C. perfringens Enterotoxin ELISA Kit) using 73 human norovirus-positive fecal samples from gastroenteritis patients across 22 episodes in Japan. RESULTS: CPE was detected in 39/73 samples using the RPLA method; however, ELISA-based examination of 10 RPLA-positive samples produced negative results. Moreover, cpe was not detected in any of the RPLA-positive (n = 32) or -negative (n = 5) samples, and C. perfringens was only isolated from one RPLA-positive sample. CONCLUSIONS: An ELISA-based testing approach may be more reliable than RPLA assays for CPE detection from human fecal samples. These findings may also be applicable to the detection of other foodborne diseases.

Evaluation of a commercial latex agglutination test for detecting rotavirus A and human adenovirus in children's stool specimens.

OBJECTIVES: Rotavirus A and human adenovirus are the two most common causes of infantile diarrhea; thus, it is of great importance to find out a rapid and accurate diagnostic method. This study aimed to evaluate the diagnostic significance of latex agglutination test for detection of rotavirus A and human adenovirus. METHODS: A prospective study was conducted on 214 diarrhea children from September 2018 to March 2019 in our hospital. Fresh stool samples were collected for detection of rotavirus A and human adenovirus by latex agglutination test and quantitative reverse transcription polymerase chain reaction (RT-qPCR). Then, the consistency of results detected by these two methods was analyzed． RESULTS: With performing the latex agglutination test, it was revealed that positive rates for detecting rotavirus A virus and human adenovirus were 23.83% (51/214) and 25.24% (54/214), respectively. Meanwhile, results of RT-qPCR showed that positive rates for detecting rotavirus A virus and human adenovirus were 58 (27.10%) and 59 (27.57%), respectively. Using RT-qPCR as the gold standard, the sensitivity and specificity of the latex agglutination test for detecting rotavirus A were 81.03% and 97.44%, and the corresponding values for detecting human adenovirus were 76.27% and 94.19%, respectively. CONCLUSION: This latex agglutination test showed a satisfactory consistency with RT-qPCR for detecting rotavirus A and human adenovirus. The mentioned commercial assay may be highly appropriate for rapid screening of rotavirus A and human adenovirus.

Comparison of the specificity of rheumatoid factor detected by latex fixation with that of regulatory rheumatoid factor.

BACKGROUND: Rheumatoid factor (RF), originally defined as pathological autoantibodies to IgG that are detected in rheumatoid arthritis, turned out to be multi-specific antibodies, some of which exhibit immunoregulatory properties. Recently, we identified a RF, the production of which confers resistance to experimental autoimmune diseases and is associated with the remission of autoimmune diseases. To differentiate the RF, we discovered from the one associated with rheumatic disease onset or progression and to reflect its immunoregulatory properties, we named it regulatory rheumatoid factor (regRF). Immunization with conformers of Fc fragments that expose regRF neoepitopes reduces collagen-induced arthritis in rats. Certain information about the specificity of classical RF and regRF indicates that these populations may be one and the same. Therefore, the aim of this study was to determine whether there is a difference between the classical RF and regRF. METHODS: Classical RF was measured in diseased blood by the latex fixation method, and regRF was detected by the agglutination of human IgG-loaded tanned erythrocytes. Competitive analysis was used to determine the specificity of rheumatoid factors. RESULTS: It was found that regRF and pathology-associated RF constitute different antibody populations. Pathology-associated RF is specific for lyophilized IgG. RegRF does not interact with IgG. RegRF is specific to conformers of IgG Fc fragments that have a reduced hinge. In latex-positive rheumatoid arthritis sera, regRF may be present in addition to pathology-associated RF. The latex fixation method detects both rheumatoid factor populations. CONCLUSION: RegRF and classical pathology-associated RF have different specificity.

Evaluation of new algorithm using TPLA as an initial syphilis screening test.

We retrospectively compared the performance of an existing syphilis diagnostic algorithm with a new algorithm that analyzes the results of Treponema pallidum latex agglutination (TPLA) tests. Of the 100 clinical blood samples, 51 were classified as positive through both Mediace TPLA and ESPLINE TP; 2/51 were classified as negative by Architect Syphilis TP, whereas 1/51 was negative as per LUMIPULSE Presto TP. The false positive rate when the results of Mediace TPLA and ESPLINE TP were combined was 1.96% versus 0% for both Architect Syphilis TP and LUMIPULSE Presto TP. The sensitivity of Mediace TPLA (98%) was comparable to that of Architect Syphilis TP (98%) but lower than that of LUMIPULSE Presto TP (100%). The specificity of Mediace TPLA was 98.0% versus 100% for Architect Syphilis TP, and versus 100% for LUMIPULSE Presto TP. We conclude that the performance of Mediace TPLA in combination with a reverse algorithm is nearly equal to that of enzyme immunoassay (EIA) or chemiluminescence immunoassay (CIA). Because TPLA is low cost, highly sensitive method for IgM detection, and is easy to operate, we have recommended its adoption for initial syphilis screening tests.

Comparison of performance characteristics between high-performance liquid chromatography and latex agglutination turbidimetric immunoassay for therapeutic drug monitoring of zonisamide.

BACKGROUND: Recently, the Nanopia® TDM Zonisamide reagent using the latex particle-enhanced turbidimetric immunoassay (LTIA) method was developed. The aim of this study was to compare the differences in serum zonisamide (ZNS) concentrations quantified by the high-performance liquid chromatography (HPLC) method and the LTIA method using a TBA-25FR analyzer. METHODS: A total of 78 samples from 33 patients were quantified by both HPLC and LTIA methods. Deproteinization was used as pretreatment for the HPLC method. The ZNS concentrations quantified by two methods were compared. RESULTS: The HPLC method had intra- and inter-day precision lower than 1.86% and 9.00%, and accuracy better than 2.44% and 6.33%, respectively. The LTIA method showed intra- and inter-day precision lower than 2.50% and 5.20%, and accuracy better than 15.80% and 10.60%, respectively. The lower limits of quantification for the HPLC and LTIA methods were 1.0 and 5.0 µg/mL, respectively. The ZNS concentration quantified by the HPLC method correlated strongly with that by the LTIA method (r = 0.953, P < 0.001). A Bland-Altman plot suggested no systematic error between ZNS concentrations quantified by HPLC and LTIA methods. CONCLUSION: This study confirmed no differences between the concentrations quantified by the HPLC and LTIA methods at both high and low concentrations, demonstrating the confidence of measurement by the LTIA method.

Diagnostic Accuracy of Latex Agglutination Turbidimetric Immunoassay in Screening Adolescents for Helicobacter pylori Infection in Japan.

BACKGROUND/AIMS: Serologic tests are commonly used for screening Helicobacter pylori infection because they not only provide quick results but also are inexpensive. A new latex agglutination serum antibody assay (LZ test) has been developed and it is expected to be as effective as conventional assays. This study aimed to calculate a reliable cutoff value for the LZ test and to estimate the sensitivity and specificity of the cutoff value in screening adolescents for H. pylori infection in Japan. METHODS: We screened junior high school students in Akita Prefecture, Japan, for H. pylori infection. We used the data of 213 such students who underwent H. pylori stool antigen (HpSA) tests in 2016. The students who had positive results with HpSA tests were diagnosed with H. pylori infection. Of the 213 students, 209 underwent the LZ test. RESULTS: The prevalence rate of H. pylori infection was 3.8% (8/209). The area under the curve for the LZ test was 0.88. The cutoff value of the LZ test was determined to be 3.1 U/mL. At this value, the sensitivity and specificity were 87.5 and 91.5%, respectively. CONCLUSION: The accuracy of the LZ test in adolescents was well balanced for sensitivity and specificity as well as for tolerable results.

Development and Evaluation of a Latex Agglutination Test for the Identification of Francisella tularensis Subspecies Pathogenic for Human.

Francisella tularensis are highly infectious bacteria causing a zoonotic disease called tularemia. Identification of this bacterium is based on antigen detection or PCR. The paper presents a latex agglutination test (LAT) for rapid identification of clinically relevant F. tularensis subspecies. The test can be performed within three minutes with live or inactivated bacteria. The possibility to test the inactivated samples reduces the risk of laboratory acquired infection and allows performing the test under BSL-2 conditions.

Epidemiology, clinical profile and role of rapid tests in the diagnosis of acute bacterial meningitis in children (aged 1-59 months).

OBJECTIVES: To study the epidemiology, clinical profile, and the role of rapid tests in the diagnosis of acute bacterial meningitis (ABM) in children (1-59 months). MATERIALS AND METHODS: A total of 250 cerebrospinal fluid (CSF) and 187 blood samples received from clinically suspected cases of ABM were processed based on standard microbiological protocols. CSF samples were also subjected to antigen and nucleic acid detection. Antibiotic susceptibility testing was done according to the Clinical Laboratory Standards Institute guidelines. Children were also evaluated for outcomes and were followed up until 6 months after discharge. RESULTS: Eighty one cases were reported to be having clinically confirmed ABM, out of which group B Streptococcus was the most common pathogen detected in 49.3% (40) patients followed by Streptococcus pneumoniae, Staphylococcus aureus, Hemophilus influenzae type b, Escherichia coli, Klebsiella pneumoniae, and Neisseria meningitidis ACYW135 in 23.4% (19), 7.4% (6), 6.1% (5), 6.1% (5), 6.1% (5), and in 1.2% (1) patients, respectively. Complications were observed in 54.3% of the children. A follow-up of 6 months after discharge was possible in 39.5% (32) patients among whom sequelae were recorded in 93.7% (30) patients. CONCLUSION: ABM remains a major cause of neurological sequelae worldwide. Although culture is the gold standard test for its detection, the investigation takes a longer time and the results are influenced by prior antimicrobial therapy. In such cases, rapid tests aid in the early diagnosis of ABM for instituting appropriate management.

Detection of gastric atrophy by circulating pepsinogens: A comparison of three assays.

BACKGROUND: Circulating levels of pepsinogens have been used in high gastric cancer-risk Asian and European populations to triage endoscopic evaluation for more severe pathology. There are different analytic methods with uncertain correlations. We therefore compared diagnostic performance of three commonly used pepsinogen assays to detect histologically confirmed gastric atrophy. METHODS: We tested plasma samples from adult patients with (n=50) and without (n=755) moderate or severe gastric corpus atrophy, as determined histologically by consensus of three expert pathologists. A single laboratory measured pepsinogens I (PgI) and II (PgII) using commercially available assays: two ELISA assays produced by Biohit (Finland) and Vector Best (Russia), and a latex agglutination assay from Eiken (Japan). Quantitative correlations were assessed by Spearman statistics. Receiver operating characteristic (ROC) curves vs histological diagnosis were calculated using both the manufacturers' and optimized cutoffs. RESULTS: Pepsinogen levels were highly correlated among the assays (pairwise Rhos: PgI≥0.84, PgII≥0.87; all P-values<.01). Based on manufacturers' cutoffs, sensitivities, specificities and areas under the ROC curve for detecting moderate to severe histological corpus atrophy by PgI/PgII were 44%/91%/0.70, 56%/84%/0.76, and 52%/90%/0.77 for Biohit, Vector Best and Eiken, respectively. Cutoffs optimized by ROC or data mining analyses did not substantially improve test performance. CONCLUSIONS: Commercial assays for pepsinogen have good relative agreement but are imperfect tests for clinical diagnosis of gastric atrophy. IMPACT: Pepsinogen testing alone does not provide sufficient information for gastric cancer risk stratification. Future investigations should focus on other potential markers, in combination with pepsinogens.

Determination of immune status in dogs against CPV-2 by recombinant protein based latex agglutination test.

Canine parvoviral enteritis is a highly contagious viral illness caused by canine parvovirus-2 (CPV-2) which affects puppies of mainly 6-20 weeks of age. Vaccination is pivotal in preventing and controlling CPV-2 infection. Determination of antibody status is a critical determinant for successful vaccination. The hemagglutination inhibition (HI) test is 'gold standard' test for quantification of antibodies specific to CPV-2, although the execution of this test is not feasible under field conditions. The present study was undertaken to develop a point of care testing to determine immune status prior to CPV-2 vaccination or to detect seroconversion in immunized dogs by latex agglutination test (LAT) using recombinant antigen. Truncated portion of VP2 protein (tVP2) of CPV-2 was selected on the basis of antigenic indices, overexpressed the recombinant protein in E. coli system and was subsequently used in development of LAT. A total of 59 serum samples obtained from vaccinated (n = 54) and healthy unvaccinated (n = 5) dogs were tested. The positivity was observed in 85% (46/54) of these dogs with varying agglutination pattern. The overall sensitivity and specificity of latex agglutination test in comparison to HI test was recorded as 90% and 88% respectively with an agreement value of 90% (CI = 95%).

Cross-reactivity in Cryptococcus antigen latex agglutination test in two commercial kits.

This article presents an examination of the cross-reactivity of pathogenic fungi with Cryptococcus neoformans in two commercial Cryptococcus antigen latex agglutination tests performed across 39 fungal strains. Some fungi were newly indicated as Cryptococcus cross-reactive, and the two kits showed differences in cross-reactive fungi.

Development and Evaluation of a New Creatine Kinase MB Mass Determination Assay Using a Latex Agglutination Turbidimetric Immunoassay with an Automated Analyzer.

BACKGROUND: The diagnosis of myocardial infraction (MI) in patients presenting to the emergency department represents a clinical challenge. It is known that creatine kinase-MB isoenzyme (CK-MB) is present in soluble cell fractions of cardiac muscle, and injury to those cells results in an increase of CK-MB in the blood. Therefore, CK-MB is a suitable clinical biomarker of myocardial infraction. METHODS: To measure CK-MB mass rapidly and easily, we developed the new reagent 'L-type Wako CK-MB mass' (L-CK-MB mass) for the latex agglutination turbidimetric immunoassay method. Using a Hitachi LABOSPECT 008, we evaluated the performance of this assay as a method for quantifying CK-MB mass, and we compared the measurement of the serum CK-MB mass concentration with this assay to that obtained using an electrochemiluminescence immunoassay (ECLIA). RESULTS: A dilution test showed linearity from 5 µg/L to 190 µg/L, and the limit of quantification of the L-CK-MB mass assay was 3.0 µg/L. The within-run CV and between-day CV were 1.0 - 4.5% and 1.8 - 4.4%, respectively. Serum CK-MB mass concentration determined using the L-CK-MB mass assay was reliably and strongly correlated with that determined using ECLIA (n = 163, r = 0.999, y = 0.977x + 0.307). CONCLUSIONS: The L-CK-MB mass assay is able to specifically determine CK-MB mass and is a very useful method for the accurate measurement of CK-MB mass for routine clinical analyses.

Pitfalls in Serological Diagnosis of Cryptococcus gattii Infections.

The detection of cryptococcal antigen by latex agglutination tests (LATs), enzyme-linked immunoassays (ELISA), or lateral flow assay (LFA) is an important tool for diagnosis of a Cryptococcus infection. Cerebrospinal fluid and/or serum samples of 10 patients with cryptococcosis due to Cryptococcus gattii or a hybrid of Cryptococcus neoformans and C. gattii were examined by three LATs (the IMMY Latex-Crypto(®) test, the Pastorex(TM) Crypto Plus, and the Remel Cryptococcus Antigen Test Kit) and the LFA made by Immuno-Mycologics. LATs based on monoclonal antibodies (mAbs) like the Pastorex(TM) Crypto Plus or the Remel Cryptococcus Antigen Test Kit turned out to have an insufficient sensitivity to detect four out of 10 C. gattii infections, including one infection by a hybrid between C. gattii and C. neoformans. Reflecting the ongoing expansion of C. gattii in geographical zones outside of tropical and subtropical areas like Mediterranean countries, Vancouver Island (British Columbia, Canada) and the Pacific Northwest region (USA), these findings are alarming because of the risk of delayed diagnosis of infections caused by C. gattii. Therefore, the preliminary serological screening for cryptococcal antigen in the case of a suspected Cryptococcus infection should be performed by using an assay with a broad range specificity and sensitivity for C. neoformans and C. gattii, including their hybrids.

Evaluation of new automated syphilis test reagents 'IMMUNOTICLES AUTO3' series: performance, biochemical reactivity, and clinical significance.

BACKGROUND: Automated nontreponemal and treponemal test reagents based on the latex agglutination method (immunoticles auto3 RPR: ITA3RPR and immunoticles auto3 TP: ITA3TP) have been developed to improve the issues of conventional manual methods such as their subjectivity, a massive amount of assays, and so on. We evaluated these reagents in regards to their performance, reactivity to antibody isotype, and their clinical significance. METHODS: ITA3RPR and ITA3TP were measured using a clinical chemistry analyzer. Reactivity to antibody isotype was examined by gel filtration analysis. RESULTS: ITA3RPR and ITA3TP showed reactivity to both IgM- and IgG-class antibodies and detected early infections. ITA3RPR was verified to show a higher reactivity to IgM-class antibodies than the conventional methods. ITA3RPR correlated with VDRL in the high titer range, and measurement values decreased with treatment. ITA3RPR showed a negative result earlier after treatment than conventional methods. ITA3TP showed high specificity and did not give any false-negative reaction. Significant differences in the measurement values of ITA3RPR between the infective and previous group were verified. CONCLUSIONS: The double test of ITA3RPR and ITA3TP enables efficient and objective judgment for syphilis diagnosis and treatments, achieving clinical availability.

Etiology and risk factors of meningitis in patients admitted at a Central Hospital in Harare.

Objective: To determine etiology and risk factors of meningitis in patients admitted a tertiary referral Hospital in Harare. Design: Cross-Sectional Study. Setting: Urban Referral Health Facility. Subjects: Patients suspected of having Meningitis admitted at Parirenyatwa Hospital were consecutively consented and recruited into the study until sample size accrual. Main Outcome Measures: Prevalence of pathogens associated with Meningitis. Risk factors of meningitis. Results: Two Hundred and Ninety Six (296) clinically suspected meningitis patients were recruited into the study, 51.7 %( n=115) were male. Meningitis was confirmed in 20.6% (n=61) cases with the following pathogen proportions, C. neoformans - 45.9 %( n=28), S. pneumoniae ­ 27.9 % (n=17), TBM ­ 4.9 %( n=3), probable viral meningitis ­ 6.6% (n=4 and other bacteria- 14.8% (n=9). Patients from crowded households were also more likely to suffer from meningitis than those from sparsely populated households (p<0.001). Conclusion: The use of Latex agglutination increases the proportion of detected pathogens both fungal and bacterial when used in conjunction with CSF gram stain and culture. Cryptococcus neoformans and S. pneumoniae are the leading causes of meningitis in patients admitted at Parirenyatwa Hospital.

[Evaluation of the Fundamental Performance of 4 Latex Agglutination Reagents to Measure Anti-TP Antibody Concentration and Detailed Investigation of Decision-Mismatched Samples].

Serological diagnosis of syphilis can be made by using the serological test for syphilis (STS) method for detecting a lipid antibody and Treponema pallidum (TP) method for detecting the anti-TP-specific antibody. In STS and TP methods, the basis using latex agglutination reaction has been used in many facilities. However, in latex agglutination, false-positive results due to non-specific reaction have sometimes been obtained in reactions of a routine laboratory test reagent detecting the anti-TP antibody used in our medical laboratory. We evaluated the fundamental performance of 4 reagents to measure anti-TP antibody concentration using latex agglutination: Reagents A, B, C and D produced by SEKISUI MEDICAL, FUJI REBIO, DENKA SEIKEN and SHINO TEST, respectively. We examined the correlations between Reagent A (routine laboratory test reagent) and Reagents B, C, and D in sera from 68 patients, and we performed additional investigation by using a neutralization test, immunochromatography, Western blotting, FTA-ABS (IgG), and STS method by an automatic analyzer for 13 decision-mismatched samples. The fundamental performance of each reagent was as good as that previously reported. Eight of the 13 decision-mismatched samples were false positives due to non-specific reaction of Reagent A. In latex agglutination non-specific reaction is inevitable. However, this study strongly suggests that using a neutralization test and immunochromatography that can be performed quickly is sufficient to verify whether positive reactions are true or false.

[Factors Affecting Non-Specific Reaction in Latex Agglutination Turbidimetric Immunoassay for Detection of Matrix Metalloproteinase-3].

PURPOSE: Measurement of matrix metalloproteinase-3(MMP-3), a marker for rheumatoid arthritis, by means of latex agglutination turbidimetric immunoassay (LTIA), has come to replace ELISA kit, but pre-improvement LTIA kit S(Sekisui Medical) frequently shows false values resulting from non-specific reactions. We analyzed factors influencing the frequency of non-specific reactions for MMP-3 detection using various methods. METHOD: Serum MMP-3 levels were measured in 1,214 routine samples or 57 panels with various immunoserological abnormalities by means of pre-improvement and improved kit S, LTIA kit E (Eiken Chemical) and ELISA kit. Non-specific reaction samples were selected either from the routine samples based on the results of correlativity tests between the kits and of dilution linearity tests, or from 57 panels with immunoserological abnormalities based on the results of recovery tests. To explore the factors causing non- specific reactions, titers of rheumatoid factor (RF), immunoglobulins and four heterophil antibodies were measured in these samples. In addition, changes in findings as a result of reduction treatment or IgM absorption were examined in selected samples to clarify the effect of heterophil antibodies. RESULTS: Samples which showed suspected false values probably due to non-specific reaction numbered 75, 15, 5 and 17 as measured with pre-improvement kit S, improved kit S, kit E and ELISA kit, respectively. Kit S also showed high rate of deviation in recovery tests on panels with immunoserological abnormalities. Rate of RF titer in samples with non-specific reactions was higher than that of other factors. Non-specific reaction could be inhibited in two samples as a result of reduction treatment or absorption of IgM subtype. CONCLUSION: Pre-improvement kit S showed a higher rate of non-specific reactions than other kits. The most common cause of non-specific reactions is thought to be high RF titer, but high levels of IgM, including heterophil antibodies, may also affect the values of MMP-3.