Development and clinical evaluation of a quantitative fluorescent immunoassay for detecting canine CRP.

Canine C-reactive protein (cCRP) is one of the major positive acute phase proteins in dogs and is commonly measured to detect and monitor systemic inflammation as well as the efficacy of treatment. Traditional methods for testing cCPR, including enzyme-linked immunosorbent assay (ELISA), have some drawbacks, such as a long time for diagnosis and the requirement of well-equipped laboratories. Therefore, there is a need for a rapid and precise diagnostic test for cCRP at point-of-care. This study assessed the accuracy, precision, and validated clinical effectiveness of a diagnostic test based on fluorescent lateral flow immunoassay to detect cCRP. For the standard cCRP concentration ranging from 0 to 200 µg/mL, the cCRP diagnostic test showed strong linearity with R2 of 0.9977 (p < 0.001), and both inter- and intra-assay CVs were <14%. The limit of detection and limit of quantitation were found to be 4.0 µg/mL and 5.0 µg/mL, respectively. The cCRP serum concentration was evaluated in 21 client-owned dogs and the results were compared to a previously validated ELISA. The Pearson Correlation Coefficient between the diagnostic test kit and ELISA was 0.942 [95% confidence interval: 0.859 to 0.976, p < 0.001], and the Bland-Altman plot indicated a bias of 26.82% [95% limits of agreement: -56.03 to 109.67], indicating a significant correlation and the agreement between the data from the cCRP diagnostic test and ELISA. In conclusion, the fluorescent immunoassay based diagnostic test is a suitable option for rapidly and precisely detecting cCRP in dogs, providing a convenient alternative to traditional methods for diagnosing acute inflammation.

Development and Clinical Evaluation of an Immunochromatography-Based Rapid Antigen Test (GenBody™ COVAG025) for COVID-19 Diagnosis.

Antigen tests for SARS-CoV-2 diagnosis are simpler and faster than their molecular counterparts. Clinical validation of such tests is a prerequisite before their field applications. We developed and clinically evaluated an immunochromatographic immunoassay, GenBody™ COVAG025, for the rapid detection of SARS-CoV-2 nucleocapsid (NP) antigen in two different clinical studies. Retrospectively, 130 residual nasopharyngeal swabs transferred in viral transport medium (VTM), pre-examined for COVID-19 through emergency use authorization (EUA)-approved real-time RT-PCR assay and tested with GenBody™ COVAG025, revealed a sensitivity and specificity of 90.00% (27/30; 95% CI: 73.47% to 97.89%) and 98.00% (98/100; 95% CI: 92.96% to 99.76%), respectively, fulfilling WHO guidelines. Subsequently, the prospective examination of 200 symptomatic and asymptomatic nasopharyngeal swabs, collected on site and tested with GenBody™ COVAG025 and EUA-approved real-time RT-PCR assay simultaneously, revealed a significantly higher sensitivity and specificity of 94.00% (94/100; 95% CI: 87.40% to 97.77%) and 100.00% (100/100; 95% CI: 96.38% to 100.00%), respectively. Clinical sensitivity and specificity were significantly high for samples with Ct values ≤ 30 as well as within 3 days of symptom onset, justifying its dependency on the viral load. Thus, it is assumed this can help with the accurate diagnosis and timely isolation and treatment of patients with COVID-19, contributing to better control of the global pandemic.

ß-arrestin2 Affects Cardiac Progenitor Cell Survival through Cell Mobility and Tube Formation in Severe Hypoxia.

BACKGROUND AND OBJECTIVES: ß-arrestin2 (ß-arr2) basically regulates multiple signaling pathways in mammalian cells by desensitization and internalization of G-protein coupled receptors (GPCRs). We investigated impacts of ß-arr2 on survival, mobility, and tube formation of cardiac progenitor cells (CPCs) obtained from wild-type (WT) mouse (CPC-WT), and ß-arr2 knock-out (KO) mouse (CPC-KO) cultured in presence or absence of serum and oxygen as non-canonical roles in GPCR system. METHODS: CPCs were cultured in Dulbecco's Modified Eagle Medium/Nutrient Mixture F-12 -based media containing fetal bovine serum and growth factors. Survival of 2 types of CPCs in hypoxia and/or serum deprivation was measured by fluorescence-activated cell sorting. Wound healing ability, and tube formation ability on Matrigel of 2 kinds of CPCs were compared in normoxic and hypoxic cultures. Protein expression related to survival and mobility were measured with the Western blot for each culture conditions. RESULT: CPC-KO showed significantly worse mobility in the wound healing assay and in tube formation on Matrigel especially in hypoxic culture than did the CPC-WT. Also, CPC-KO showed significantly higher apoptosis fraction in both normoxic and hypoxic cultures than did the CPC-WT. Expression of proteins associated with cell survival and mobility, e.g., protein kinase B (Akt), ß-catenin, and glycogen synthase kinase-3ß (GSK-3ß) was significantly worse in CPC-KO. CONCLUSIONS: The CPC-KO had significantly worse cell mobility, tube formation ability, and survival than the CPC-WT, especially in the hypoxic cultures. Apparently, ß-arr2 is important on CPC survival by means of mobility and tube formation in myocardial ischemia.