Evaluation of the analytical and clinical performance of a new high-sensitivity cardiac troponin I assay: hs-cTnI (CLIA) assay.

OBJECTIVES: Cardiac troponin (cTn) is the key biomarker for diagnosis of acute coronary syndrome (ACS). We performed a complete assessment of the high-sensitivity cardiac troponin I (hs-cTnI) (CLIA) assay on the analytical performance and clinical diagnostic performance, which was compared with Abbott ARCHITECT hs-cTnI assay. METHODS: Sex-specific 99th percentile upper reference limits (URLs) were determined from a healthy population of 424 males and 408 females. High-sensitivity performance was assessed by examining the imprecision at sex-specific URLs and the detectable results above LoD in a cohort of healthy population. The diagnostic performance of the hs-cTnI (CLIA) assay was validated in a population of 934 patients with suspected ACS. RESULTS: The 99th percentile URLs were 15.3 ng/L for female, 31.3 ng/L for male and 24.2 ng/L for overall population. The total imprecision near the sex-specific 99th percentile URLs were <5 %. 76.74 % of females, 97.12 % of males and 86.69 % of overall population had cTnI values exceeding the LoD, which met the criteria of high-sensitivity troponin assay. No cross-reactivity or interference was identified. The diagnostic sensitivity, specificity, PPV, NPV, and AUC of hs-cTnI (CLIA) assay were 97.97 , 90.70, 79.02, 99.21 % and 0.9885, respectively, which were comparable to ARCHITECT hs-cTnI assay. CONCLUSIONS: hs-cTnI (CLIA) assay is a high-sensitivity troponin I method with high precision, sensitivity and specificity. The clinical diagnostic performance of hs-cTnI (CLIA) is comparable to the established ARCHITECT hs-cTnI assay. Mindray's hs-cTnI (CLIA) assay is an attractive alternative for diagnosis of myocardial infarction with a high level of accuracy and safety.

A comparison of the chemiluminescence immunoassay and Crithidia luciliae immunofluorescence test in detecting anti-dsDNA antibodies and assessing the activity of systemic lupus erythematosus.

OBJECTIVES: This study aimed to compare the YHLO chemiluminescence immunoassay (CLIA) with the Crithidia luciliae immunofluorescence test (CLIFT) to detect anti-dsDNA antibodies and its correlation with disease activity in systemic lupus erythematosus (SLE). METHOD: In total, 208 patients diagnosed with SLE, 110 other autoimmune patients, 70 infectious disorders patients, and 105 healthy people were enrolled in this study. Serum samples were tested using CLIA in a YHLO chemiluminescence system and CLIFT. RESULTS: The overall agreement between YHLO CLIA and CLIFT was 76.9% (160/208), with a moderate correlation (kappa = 0.530, p < 0.001). The sensitivity of YHLO CLIA and CLIFT were 58.2% and 55.3%, respectively. The specificity of YHLO CLIA and CLIFT were 95.1% and 99.3%, respectively. The sensitivity of YHLO CLIA was increased to 66.8% with a specificity of 93.6% when the cut-off value was set at 24 IU/mL. Spearman's correlation coefficient between the quantitative results of YHLO CLIA and the titers of CLIFT was 0.59 (p < .01). A significant correlation was found between the anti-dsDNA results detected by YHLO CLIA and the SLE Disease Activity Index 2000 (SLEDAI-2K). Spearman's correlation coefficient between YHLO CLIA and SLEDAI-2K (r = 0.66, p < .01) was higher than that of CLIFT (r = 0.60, p < .01). CONCLUSIONS: Good correlation and agreement were found between YHLO CLIA and CLIFT. In addition, there was a significant correlation between YHLO CLIA and the SLE Disease Activity Index, which was superior to that of CLIFT. The YHLO chemiluminescence system is recommended for the assessment of disease activity.

Analytical evaluation of a GAD65 antibodies chemiluminescence immunoassay for CSF in neurological syndromes.

OBJECTIVES: Antibodies against glutamic acid decarboxylase isoform 65 (GAD-Ab) have been found in different severe neurological conditions associated with altered synthesis of γ-aminobutyric acid (GABA). Serum GAD-Ab can be found in up to 90 % of patients with type 1 diabetes mellitus (T1DM), mostly at relatively low concentrations, while high concentrations of GAD-ab are thought to be more frequently associate to a neurological condition, with levels 100-folds higher than those found in T1DM. Although CSF testing is recommended when suspecting a GAD-associated neurological syndrome, no commercial immunoassay is validated for this use and no cut-off is internationally recognized to support the diagnosis. METHODS: In this study we validated CSF testing of GAD-Ab on an automated chemiluminescence (CLIA) immunoassay that had previously shown good agreement with ELISA on serum. RESULTS: We tested 43 CSF from patients with typical GAD-associated neurological disorders and patients with other neurological conditions, identifying a clinical cut-off of 18 kIU/L that discriminated GAD-disease with an area under the curve (AUC) of 0.921. CLIA showed good analytical performances on repeatability and recovery tests in CSF and confirmed an excellent agreement with ELISA. CONCLUSIONS: GAD-Ab associated neurological disorders are rare but CSF testing for GAD-Ab is a common request for neurologists when suspecting an insidious autoimmune central nervous system disease. CLIA platforms are expected to be increasingly adopted in clinical laboratories due to their flexibility and reliability, therefore studies on decisional levels should be implemented for improving the interpretation and utilization of laboratory data.

Chemiluminescence Immunoassay Method of Urinary Liver Fatty-acid-binding Protein as a Promising Candidate for Kidney Disease.

Liver fatty acid binding protein (L-FABP) is an intercellular lipid chaperone protein that selectively combines with unsaturated free fatty acids and transports them to mitochondria or peroxisomes. L-FABP is a promising biomarker for the early detection of renal diseases in humans. Herein a chemiluminescence method (CLIA) was demonstrated to measure the level of urinary L-FABP in the urinary samples. An anti-(L-FABP)-magnetic beads complex was prepared to capture the analyte target. Sensitivity, precision, accuracy, interference effect, high-dose hook effect of the developed assay were evaluated. Under the suitable experimental parameters, the established method have a wide linear range (0.01-10 ng/mL) and also showed a sufficiently low limit of detection of 0.0060 ng/mL. Besides, the satisfactory recoveries of the method in the urinary were ranged from 97.74%-112.32%, which was well within the requirement of clinical analysis. Furthermore, this proposed method has been successfully applied to the clinical determination of L-FABP in patients who have been diagnosed with kidney disease. The results showed that CLIA could accurately and rapidly determine the urinary level of L-FABP with high-throughput, which could be useful as a new tool to predict complications in patients with kidney disease. The clinical trial was approved by Shuyang Hospital of Traditional Chinese Medicine Ethics Committee: 20,210,202-001 at February 2, 2021.

Automated detection of neutralizing SARS-CoV-2 antibodies in minutes using a competitive chemiluminescence immunoassay.

The SARS-CoV-2 pandemic has shown the importance of rapid and comprehensive diagnostic tools. While there are numerous rapid antigen tests available, rapid serological assays for the detection of neutralizing antibodies are and will be needed to determine not only the amount of antibodies formed after infection or vaccination but also their neutralizing potential, preventing the cell entry of SARS-CoV-2. Current active-virus neutralization assays require biosafety level 3 facilities, while virus-free surrogate assays are more versatile in applications, but still take typically several hours until results are available. To overcome these disadvantages, we developed a competitive chemiluminescence immunoassay that enables the detection of neutralizing SARS-CoV-2 antibodies within 7 min. The neutralizing antibodies bind to the viral receptor binding domain (RBD) and inhibit the binding to the human angiotensin-converting enzyme 2 (ACE2) receptor. This competitive binding inhibition test was characterized with a set of 80 samples, which could all be classified correctly. The assay results favorably compare to those obtained with a more time-intensive ELISA-based neutralization test and a commercial surrogate neutralization assay. Our test could further be used to detect individuals with a high total IgG antibody titer, but only a low neutralizing titer, as well as for monitoring neutralizing antibodies after vaccinations. This effective performance in SARS-CoV-2 seromonitoring delineates the potential for the test to be adapted to other diseases in the future.

Evaluation of chemiluminescence immunoassay as a screening test for syphilis on blood donor samples.

BACKGROUND AND OBJECTIVE: Global re-emergence of syphilis among blood donors necessitates novel diagnostic and prevention approaches that encourage timely intervention. Thus, the present study was planned to evaluate the efficiency of Chemiluminescence immunoassay (CLIA) as a screening test for syphilis. MATERIAL AND METHODS: This prospective cross-sectional observational study was conducted from October 2021 to September 2022. A total of 344 donors were enrolled by purposive sampling method, including additional 16 donors who were reactive by the Rapid plasma reagin test (RPR) during the study period. Data from three screening tests - RPR test, Treponema pallidum haemagglutination assay (TPHA) and CLIA for 360 blood donors were analysed. TPHA was considered the gold standard test. RESULTS: Of the total 360 samples tested, 21 (5.8 %) were reactive by the RPR test. Of these 21 RPR reactive samples, 19 (90.5 %) were reactive by both TPHA and CLIA, while 2 (9.5 %) RPR reactive samples were non-reactive by both TPHA and CLIA. Of the remaining 339 RPR non-reactive samples, 1 (0.3 %) sample was reactive by both TPHA and CLIA, and 1 (0.3 %) was reactive by CLIA alone. CLIA was found to have sensitivity and specificity of 100 % and 99.7 % and positive predictive value (PPV) and negative predictive values (NPV) of 95.2 % and 100 % respectively, while it was 95 %, 99.4 %, 90 %, and 99.7 %, respectively, with the RPR test. CONCLUSION: CLIA was found to have a higher sensitivity, specificity, PPV and NPV than the RPR test. Thus, CLIA can be an acceptable alternative for syphilis screening in blood donors.

One-Pot Synthesis of Enzyme and Antibody/CaHPO4 Nanoflowers for Magnetic Chemiluminescence Immunoassay of Salmonella enteritidis.

In this study, through a bioinspired strategy, the horseradish peroxidase (HRP) and antibody (Ab) were co-embedded into CaHPO4 to prepare HRP-Ab-CaHPO4 (HAC) bifunctional hybrid nanoflowers by one-pot mild coprecipitation. The as-prepared HAC hybrid nanoflowers then were utilized as the signal tag in a magnetic chemiluminescence immunoassay for application in the detection of Salmonella enteritidis (S. enteritidis). The proposed method exhibited excellent detection performance in the linear range of 10-105 CFU/mL, with the limit of detection (LOD) of 10 CFU/mL. This study indicates great potential in the sensitive detection of foodborne pathogenic bacteria in milk with this new magnetic chemiluminescence biosensing platform.

Low Risk of Serological Cross-Reactivity between the Dengue Virus and SARS-CoV-2-IgG Antibodies Using Advanced Detection Assays.

Several studies have reported serological cross-reactivity of the immune responses between SARS-CoV-2 and DENV. Most of the available studies are based on the point-of-care rapid testing kits. However, some rapid test kits have low specificity and can generate false positives. Hence, we aimed to investigate the potential serological cross-reactivity between SARS-CoV-2 and DENV-IgG antibodies using advanced assays including chemiluminescence immunoassay (CLIA) and enzyme-linked immunosorbent assay (ELISA) test. A total of 90 DENV-IgG-ELISA-positive and 90 DENV-IgG-ELISA-negative prepandemic sera were tested for anti-SARS-CoV-2-IgG using the automated CL-900i CLIA assay. Furthermore, a total of 91 SARS-CoV-2-IgG-CLIA-positive and 91 SARS-CoV-2-IgG-CLIA-negative postpandemic sera were tested for anti-DENV-IgG using the NovaLisa ELISA kit. The DENV-IgG-positive sera resulted in five positives and 85 negatives for SARS-CoV-2-IgG. Similarly, the DENV-IgG-negative sera also resulted in 5 positives and 85 negatives for SARS-CoV-2-IgG. No statistically significant difference in specificity between the DENV-IgG-positive and DENV-IgG-negative sera was found (p value = 1.00). The SARS-CoV-2-IgG-positive sera displayed 43 positives, 47 negatives, and 1 equivocal for DENV-IgG, whereas the SARS-CoV-2-IgG-negative sera resulted in 50 positives, 40 negatives, and 1 equivocal for DENV-IgG. No statistically significant difference in the proportion that is DENV-IgG positive between the SARS-CoV-2-IgG-positive and SARS-CoV-2-IgG-negative sera (p value = 0.58). In conclusion, there is a low risk of serological cross-reactivity between the DENV and SARS-CoV-2-IgG antibodies when using advanced detection assays.

Preparation of a novel group of chemiluminescent N-substituted acridinium esters.

Several novel N-substituted acridinium esters 7-16 containing a 10-methyl, 10-dodecyl, or 10-(ω-[succinimidyloxycarbonyl]alkyl) group have been synthesized and their chemiluminescent properties have been tested. Their chemiluminescent efficiencies and hydrolytic stabilities have been found to be affected by the characteristics of the group on the nitrogen atom. Dibromo-substituted leaving groups slightly accelerate the chemiluminescence process.

Performance evaluation of the new Chemiluminescence Immunoassay CL-1200i Thyroid Panel.

Aim of this work was to verify the analytical performance of thyroid panel tests measured by chemiluminescence immunoassay (CLIA) CL-1200i and to validate its efficacy as laboratory test for thyroid disorder.Serum samples were obtained by standard centrifugation, thawed and assayed in a blinded fashion, and in a single batch. This study compares the values of thyroid panel tests measured by Mindray CL-1200i chemiluminescent system to the Abbott platforms for TSH, FT3, FT4, and Beckman Coulter for Tg, TgAb, and TPOAb on patient serum samples. A total of 180 randomly selected patients including both hospitalized and ambulatory patients from the Policlinico Tor Vergata (PTV) of the University of Rome Tor Vergata were used. In all analyses performed, the thyroid panel tests of the Mindray platform showed discriminative ability to quantitatively assess the analyte involved in thyroid disease and disorder. This study verified that Mindray CL-1200i chemiluminescent system thyroid panel tests is a valid method for obtaining a quantitative analysis of thyroid disorders. It showed high diagnostic efficiency and could represent a valid tool with a potential reduction in time and workload for the diagnosis.

Use of serum progesterone and prostaglandin F2α metabolite levels to predict onset of parturition in the bitch.

The prediction of time to onset of parturition in a preparturient bitch is of great clinical value, particularly for bitches at high risk of dystocia and those lacking relevant clinical data from the time of breeding. In a previous study, four cut-offs for plasma progesterone levels, measured by radioimmunoassay, were shown to be useful for predicting the likelihood of a bitch entering stage one of parturition within defined time intervals. The first aim of the current study was to evaluate these cut-offs in a clinical setting, using serum progesterone samples drawn from preparturient bitches 12-hourly instead of 6-hourly and assessed using chemiluminescence immunoassay. Furthermore, the use of 13,14-dihydro-15-keto-prostaglandin F2α, (PGFM), a metabolite of prostaglandin F2α , in predicting the time to onset of parturition was evaluated. Forty bitches carrying two or more foetuses were admitted to a specialist veterinary reproduction hospital 53 d after the onset of cytological dioestrus when that date was known, or 57 d after the last mating. Vaginal speculum examinations were performed every 6 hr until cervical dilatation was visualized (time of cervical dilatation; TCD). Serum samples were collected at 08h00 and 18h00 daily until TCD. All bitches underwent elective caesarean section at TCD. Results of this study show that approximately 5% and 10% of preparturient bitches will reach TCD within 12 hr despite a serum progesterone level of at least 15.8 nmol/L and 8.7 nmol/L respectively. In addition, there is a 95% probability that a preparturient bitch will reach TCD within 48 hr if her serum progesterone level is below 8.7 nmol/L, and a 91% probability of her reaching TCD within 24 hr if her serum progesterone level is below 3.18 nmol/L. Approximately 90% of bitches that demonstrate a 20% increase in PGFM over a 12-hr period are likely to be within 36 hr of TCD. These results provide useful benchmarks for the management of canine parturition.

Development of an Indirect Chemiluminescence Immunoassay Using a Multiepitope Recombinant Protein To Specifically Detect Antibodies against Foot-and-Mouth Disease Virus Serotype O in Swine.

Foot-and-mouth disease virus (FMDV) has led to serious losses in animal husbandry worldwide. Seromonitoring of FMDV postvaccination is important for the control and eradication of foot-and-mouth disease (FMD) in regions and countries where vaccination is widespread. However, many commercial kits present high false-positive rates. In this study, a multiepitope-based indirect chemiluminescence immunoassay (ME-CLIA) was developed for specifically detecting antibodies against FMDV serotype O in swine sera. The developed method presented high diagnostic sensitivity and excellent diagnostic specificity, and it could detect a broad spectrum of antibodies against FMDV serotype O. The diagnostic performance, accuracy rate, and analytical sensitivity of ME-CLIA were compared with those of three commercial kits. The immune protection value of multiple-epitope recombinant vaccine detected using ME-CLIA was preliminarily determined by observation of clinical symptoms postimmunization challenge, the results of which indicated that the ME-CLIA can be employed as a matching detection method for evaluating multiple-epitope recombinant vaccine. The percent positive values of ME-CLIA determined using swine vaccinated with inactivated vaccine were significantly positively correlated with the titers of liquid-phase-blocking enzyme-linked immunosorbent assay (ELISA) (LBPE) (r = 0.8361; P < 0.0001). These results indicated that ME-CLIA is suitable for detection of antibodies against FMDV serotype O in swine and for potency evaluation of multiple-epitope and inactivated vaccines.

Medium-term serostatus in Spanish case series recovered from SARS-CoV-2 infection.

The medium-term serologic response of SARS-CoV-2 infection recovered individuals is not well known. The aims were to quantify the incidence of seropositive failure in the medium term in a cohort of patients with different COVID-19 severity and to analyze its associated factors. Patients who had recovered from mild and severe forms of SARS-CoV-2 infection in an Academic Spanish hospital (March 12-May 2, 2020), were tested for total anti-SARS-CoV-2 antibodies by electrochemiluminescence immunoassay (Elecsys Anti-SARS-CoV-2 test; Roche Diagnostics GmbH). The non-seropositive status (seropositive failure) incidence (95% CI) was determined. Associations were tested by multiple logistic regression in a global cohort and severe pneumonia subpopulation. Of 435 patients with PCR-confirmed SARS-CoV-2, a serological test was carried out in 325: 210 (64.6%) had severe pneumonia (hospitalized patients), 51 (15.7%) non-severe pneumonia (managed as outpatients), and 64 (19.7%) mild cases without pneumonia. After a median (IQR) of 76 days (70-83) from symptom onset, antibody responses may not consistently develop or reach levels sufficient to be detectable by antibody tests (non-seropositive incidence) in 6.9% (95% CI, 4.4-10.6) and 20.3% (95% CI, 12.2-31.7) of patients with and without pneumonia, respectively. Baseline independent predictors of seropositive failure were higher leukocytes and fewer days of symptoms before admission, while low glomerular filtrate and fever seem associated with serologic response. Age, comorbidity or immunosuppressive therapies (corticosteroids, tocilizumab) did not influence antibody response. In the medium-term, SARS-CoV-2 seropositive failure is not infrequent in COVID-19 recovered patients. Age, comorbidity or immunosuppressive therapies did not influence antibody response.

Development of a competitive chemiluminescence immunoassay using a monoclonal antibody recognizing 3B of foot-and-mouth disease virus for the rapid detection of antibodies induced by FMDV infection.

BACKGROUND: Foot-and-mouth disease (FMD) is a devastating animal disease. Anti-non-structural protein (NSP) antibody detection is very important for confirming suspected cases, evaluating the prevalence of infection, certifying animals for trade and controlling the disease. METHODS: In this study, a competitive chemiluminescence immunoassay (3B-cCLIA) was developed for the rapid detection of antibodies against NSPs in different species of livestock animals using the monoclonal antibody (mAb) 9E2 as a competitive antibody that recognizes NSP 3B. RESULTS: The cut-off value (50%), diagnostic sensitivity (Dsn) (97.20%, 95.71%, and 96.15%) and diagnostic specificity (Dsp) (99.51%, 99.43%, and 98.36) of the assay were estimated by testing a panel of known-background sera from swine, cattle and sheep, respectively. The accuracy rate of the 3B-cCLIA was further validated and subsequently compared with that of two commercial diagnostic kits. The early diagnostic results showed that antibodies recognizing NSPs developed later (approximately 1-2 days) than antibodies recognizing structural proteins. Furthermore, anti-NSP antibody presence in animals vaccinated multiple times (false positives), especially cattle and sheep, was confirmed, and the false-positive rate increased with the number of vaccinations. CONCLUSIONS: These results indicate that the 3B-cCLIA is suitable for the rapid detection of antibodies against FMDV NSP 3B in a wide range of species.

A point-of-care chemiluminescence immunoassay for pepsinogen I enables large-scale community health screening.

Pepsinogen I (PGI) can reflect the morphology and function of the gastric mucosa. Accordingly, the large-scale community health screening of PGI can dramatically increase the early diagnosis rate of gastric cancer. However, PGI testing can only be carried out in comprehensive hospitals and health examination centers. To ameliorate this issue, a point-of-care chemiluminescent immunoassay for PGI was developed in a fully automated miniaturized instrument. This instrument was especially developed for health check-ups in the grassroots communities; its volume of which is only 0.18 m3. Critically, the entire detection process for a single sample only requires 20 min, and the samples can be loaded continuously, making the method suitable for high-throughput analysis. The assay displayed an excellent detection limit of 0.048 ng/mL with a broad detection range of 0-200 ng/mL. Furthermore, this assay exhibited high sensitivity and specificity, had low intra- and inter-assay coefficients of variation (<10%), and was not affected after storage at 37 °C for 7 days. The assay was used to detect PGI in 95 clinical serum samples, and the results were highly correlated with those that were clinically tested (correlation coefficient, R2 = 0.998). Hence, the method established in this work has great application value and can be broadly applied for the large-scale screening of gastric cancer in resource-limited areas.

Development of chemiluminescence method based on serum type I collagen hydroxyl terminal peptide ß special sequence (ß-CTX).

The objective of this work is to develop and verify the analytical performance of a chemiluminescence immunoassay for the specific sequence ß-carboxy-terminal cross-linking telopeptide of type I collagen (ß-CTX) in human serum. Two specific monoclonal antibodies (mAb-8A03 and mAb-3D12) with high affinity for ß-CTX were selected, and, under optimized conditions, a chemiluminescence immunoassay method (CLIA) for ß-CTX was established. The CLIA of ß-CTX detected ß-CTX in a wide range of 2.0-6000 ng/L. The recovery rate in serum is 95-105%, the specificity is high, and the cross-reaction rate with common easily interfering substances is low (not more than 0.01%). The CLIA correlates well with Roche electrochemiluminescence immunoassay (ECLIA), with a correlation coefficient of 0.9551, which fully meets the requirements of clinical analysis. The developed ß-CTX CLIA kit has high sensitivity and good stability. It has the same performance as the commercial Roche ECLIA kit and can be applied clinically.

A quantitative detection of Cardiotrophin-1 in chronic heart failure by chemiluminescence immunoassay.

BACKGROUND: Cardiotrophin-1 (CT-1) is a cytokine that could induce cardiomyocytes hypertrophy and dysfunction. Plasma CT-1 might serve as a cardiac biomarker both in diagnosis, staging, and prognostic assessment of heart failure. METHODS: In this study, a one-step paramagnetic particles-based chemiluminescence immunoassay (MPs-CILA) for rapid and sensitive detection of plasma CT-1 was established. Plasma samples were directly incubated with biotin-labeled anti-CT-1 antibody (bio-Ab) and acridine ester labeled anti-CT-1 antibody (AE-Ab) to form sandwiched complex. The sandwiched CT-1 was then captured by streptavidin modified paramagnetic particles (MPs-SA) for rapid separation and signal generation. RESULTS: The proposed MPs-CLIA presents a laudable linear relationship ranging from 7.8 pg/mL to 200 ng/mL with a detection limit of 1.0 pg/mL. The recoveries of spiked human plasma samples at low (10pg/mL), medium (100 pg/mL), and high (800 pg/mL) levels of CT-1 were 96%, 104%, and 110% respectively. The intra-analysis coefficient variation (CVs) of the 3 samples was 8.92%, 6.69%, and 3.54%, respectively. And the inter-analysis coefficient variation (CVs) was 9.25%, 10.9%, and 4.3%, respectively. These results strongly indicate high sensitivity, wide linear range, acceptable precision, and applicable reproducibility of the proposed method to detect plasma level of CT-1. Finally, Plasma CT-1 from 140 subjects with or without chronic heart failure was analyzed to assess the clinical application of MPs-CILA. CONCLUSIONS: Noteworthily, the MPs-CLIA method is highly automated such that it is suitable for high-throughput detection of CT-1 in clinical inspection.

Measurement of methotrexate in human cerebrospinal fluid using a chemiluminescence immunoassay intended for serum and plasma matrices.

BACKGROUND: The concentration of MTX in blood is often measured quickly and easily by immunoassays. Thus, immunoassays may facilitate the easy determination of the concentration of MTX in the cerebrospinal fluid (CSF). In this study, we measured methotrexate (MTX) concentrations in the CSF using a high-performance liquid chromatography (HPLC) method intended for analyzing CSF matrices and a chemiluminescence immunoassay (CLIA) method intended for assessing serum and plasma matrices and verified the differences in the results of the two methods. METHODS: HPLC analysis for MTX in the CSF was performed using a Prominence UFLC system with a C18 column. The HPLC method was validated in accordance with the 2018 FDA guideline. The CLIA method was performed using an ARCHITECT i1000SR system intended for serum and plasma matrices. A total of 47 CSF samples (14 clinical and 33 spiked specimens) were analyzed using the two methods. RESULTS: The HPLC method passed the validation criteria. The concentration of MTX in the same sample, determined using the HPLC and CLIA methods, differed proportionally; the percent difference in the concentrations averaged -23.0% (95% confidence interval: -36.9% to -9.1%) as revealed by the Bland-Altman plot. The relationship between the measured values, evaluated using the Passing-Bablok regression, was as follows: HPLC = 1.205 × CLIA - 0.024. CONCLUSION: The equation deduced in this study can be used to correct the concentration of MTX measured using the CLIA method.

Determination of plasma ß-amyloids by rolling circle amplification chemiluminescent immunoassay for noninvasive diagnosis of Alzheimer's disease.

A rolling circle amplification chemiluminescence immunoassay (RCA-CLIA) was developed for precise quantitation of Aß in plasma. Capture antibodies conjugated with magnetic beads and detection antibodies with collateral single-stranded DNA (ssDNA) were bound to Aß42/Aß40 antigens to form a typical double-antibody sandwich structure. The RCA reaction was triggered by the addition of ssDNA, which generated products with a large number of sites for the binding of acridinium ester (AE)-labeled detection probes, thereby realizing the purpose of the amplification. The RCA-CLIA method had higher sensitivity than conventional CLIA without loss of specificity. Under optimum conditions, the linear range of Aß42 and Aß40 detection was 3.9-140 pg/mL and 3.9-180 pg/mL, respectively, with corresponding low detection limits of 1.99 pg/mL and 3.14 pg/mL, respectively. Plasma Aß42 and Aß40 were detected in the blood of 21 AD patients and 22 healthy people, wherein this ratio could significantly distinguish AD patients from healthy individuals with a sensitivity of 90.48% and specificity of 63.64% for a cutoff value of 154. The Aß42/Aß40 ratio of plasma acts as an accurate indicator for AD diagnosis; therefore, detection of plasma Aß using the RCA-CLIA exhibits great potential in noninvasive diagnosis and progressive assessment of AD.

A Peptide-Based Magnetic Chemiluminescence Enzyme Immunoassay for Serological Diagnosis of Coronavirus Disease 2019.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a novel ß-coronavirus, causes severe pneumonia and has spread throughout the globe rapidly. The disease associated with SARS-CoV-2 infection is named coronavirus disease 2019 (COVID-19). To date, real-time reverse-transcription polymerase chain reaction (RT-PCR) is the only test able to confirm this infection. However, the accuracy of RT-PCR depends on several factors; variations in these factors might significantly lower the sensitivity of detection. METHODS: In this study, we developed a peptide-based luminescent immunoassay that detected immunoglobulin (Ig)G and IgM. The assay cutoff value was determined by evaluating the sera from healthy and infected patients for pathogens other than SARS-CoV-2. RESULTS: To evaluate assay performance, we detected IgG and IgM in the sera from confirmed patients. The positive rate of IgG and IgM was 71.4% and 57.2%, respectively. CONCLUSIONS: Therefore, combining our immunoassay with real-time RT-PCR might enhance the diagnostic accuracy of COVID-19.