Development of a time-resolved fluorescence immunoassay kit for detecting canine coronavirus and parvovirus through double labeling.

OBJECTIVE: Canine enteric coronavirus (CCV) and canine parvovirus type 2 (CPV-2) are the main pathogens responsible for acute gastroenteritis in dogs, and both single and mixed infections are common. This study aimed to establish a double-labeling time-resolved fluorescence immunoassay (TRFIA) to test and distinguish CCV and CPV-2 diseases. METHODS: A sandwich double-labeling TRFIA method was established and optimized using europium(III) (Eu3+)/samarium(III) (Sm3+) chelates. CCV/CPV-2 antigens were first captured by the immobilized antibodies. Then, combined with Eu3+/Sm3+-labeled paired antibodies, the Eu3+/Sm3+ fluorescence values were detected after dissociation to calculate the CCV/CPV-2 ratios. The performance, clinical performance and methodology used for laboratory (sensitivity, specificity, accuracy and stability) testing were evaluated. RESULTS: A double-label TRFIA for CCV and CPV-2 detection was optimized and established. The sensitivity of this TRFIA kit was 0.51 ng/mL for CCV and 0.80 ng/mL for CPV-2, with high specificity for CCV and CPV-2. All the accuracy data were less than 10%, and the recovery ranged from 101.21 to 110.28%. The kits can be temporarily stored for 20 days at 4 °C and can be stored for 12 months at temperatures less than - 20 °C. Based on a methodology comparison of 137 clinically suspected patients, there was no statistically significant difference between the TRFIA kit and the PCR method. Additionally, for CCV detection, the clinical sensitivity was 95.74%, and the clinical specificity was 93.33%. For CPV-2 detection, the clinical sensitivity was 92.86%, and the clinical specificity was 96.97%. CONCLUSION: In this study, a double-label TRFIA kit was prepared for CCV and CPV-2 detection with high laboratory sensitivity, specificity, accuracy, stability, clinical sensitivity and specificity. This kit provides a new option for screening/distinguishing between CCV and CPV-2 and may help improve strategies to prevent and control animal infectious diseases in the future.

Quantitative detection of mpox antigen using time-resolved fluorescence immunochromatography.

Recently, concern has been raised about the spread of human mpox virus, and the demand for rapid detection reagents for mpox virus has increased. This study aims to establish a time-resolved fluorescence immunochromatography (TRFICO) method for qualitative/quantitative detection of mpox virus. A double-antibody sandwich TRFICO method was optimized and established using mpox recombinant fusion antigen and its paired monoclonal antibody. The test performance of the method was evaluated using mpox fusion antigen and control serum, including sensitivity, linearity range, specificity, precision, and reference interval. We successfully established a TRFICO method for qualitative/quantitative detection of mpox antigen, its linearity range 0-100 ng/mL, analytical sensitivity 0.017 ng/mL, and reference intervals greater than 0.045 ng/mL. No cross-reaction was detected with various poxvirus and clinical negative controls, with good specificity. All average recoveries of the intra- and inter-batch ranged from 81.33% to 97.83%, and all CVs were below 10%. Additionally, the TRFICO strips can be stably stored at 37°C for 7 days without significant changes in the fluorescence intensity. This TRFICO method, with high sensitivity, linearity range, specificity, precision, and stability with 16-min detection time, provides a new option for qualitative/quantitative and convenient testing of mpox virus.

Development of a time-resolved immunochromatographic test strip for rapid and quantitative determination of GFAP in serum.

Glial fibrillary acidic protein (GFAP) in serum has been shown as a biomarker of traumatic brain injury (TBI) which is a significant global public health concern. Accurate and rapid detection of serum GFAP is critical for TBI diagnosis. In this study, a time-resolved fluorescence immunochromatographic test strip (TRFIS) was proposed for the quantitative detection of serum GFAP. This TRFIS possessed excellent linearity ranging from 0.05 to 2.5 ng/mL for the detection of serum GFAP and displayed good linearity (Y = 598723X + 797198, R2 = 0.99), with the lowest detection limit of 16 pg/mL. This TRFIS allowed for quantitative detection of serum GFAP within 15 min and showed high specificity. The intra-batch coefficient of variation (CV) and the inter-batch CV were both < 4.0%. Additionally, this TRFIS was applied to detect GFAP in the serum samples from healthy donors and patients with cerebral hemorrhage, and the results of TRFIS could efficiently discern the patients with cerebral hemorrhage from the healthy donors. Our developed TRFIS has the characteristics of high sensitivity, high accuracy, and a wide linear range and is suitable for rapid and quantitative determination of serum GFAP on-site.

Determination of serum CA724 levels using fluorescence immunochromatography.

BACKGROUND: Carbohydrate antigen 724 (CA724) is a sensitive and specific indicator for multiple malignant tumors. The aim of this study was to establish a Eu-time resolved fluorescence immunochromatography (Eu-TRFICO) method for quantitative detection of CA724 in serum. METHODS: Eu-TRFICO strips were optimized and assembled. The sensitivity, specificity and precision were evaluated using CA724 standard dilutions and matrix serum. Meanwhile, the reference interval, comparison, and sensitivity/specificity were performed using clinical negative/positive gastric cancer serum samples. RESULTS: The standard curve equation was y = 9.869 x - 154.12 (R2 = 0.993), and the sensitivity was 0.42 U/mL. The common interferents in serum could not affect the quantitative results with low cross-reactivities (all no more than 1.09%). All average recoveries of the intra- and interbatch ranged from 102.38 to 106.40%, and all CVs were below 10%. The reference interval of the healthy subjects was < 4.68 U/mL and the reference interval of the subjects with grade I/II gastric cancer was > 9.54 U/mL. Additionally, a high Pearson r (0.9503) and sensitivity/specificity (92.86%/94.20%) were obtained. CONCLUSION: This study prepared Eu-TRFICO strips with high sensitivity, specificity, precision and satisfactory clinical testing performance, which provides more options for clinical quantitative and convenient testing of CA724.

Time-resolved fluorescence immunoassay (TRFIA) for the simultaneous detection of hs-CRP and lipoprotein(a) in serum.

Elevated serum high-sensitivity C-reactive protein (hs-CRP) and lipoprotein(a) (Lp(a)) levels are associated with the development of native coronary atherosclerosis. We aimed to establish a new method for the simultaneous detection of hs-CRP and Lp(a) to predict the development of atherosclerosis. A one-step time-resolved fluorescence immunoassay (TRFIA) with europium(III) (Eu3+ ) or samarium(III) (Sm3+ ) labels was established, and the performance of this TRFIA (in terms of sensitivity, specificity, accuracy, and cutoff values) was evaluated using clinical serum samples and compared with those of registered kits. The sensitivity was 0.052 µg/ml for hs-CRP and 0.64 µg/ml for Lp(a). The intra-assay and inter-assay cross-reactivities (CVs) were very low, ranging from 2.05% to 4.67% for hs-CRP and from 2.42% to 6.43% for Lp(a). The CVs were very low (<0.34% and <2.65%, respectively) with five interferents. Additionally, there was a high Pearson coefficient between the present TRFIA method and the registered kits (R2 = 0.9967 and 0.9906, respectively). These data indicate that this study developed a TRFIA method that can be used for the quantitative detection of hs-CRP and Lp(a) in serum with high sensitivity, specificity, and accuracy. This TRFIA provides a new method for predicting the development of atherosclerosis.

Determination of human COVID-19 total antibodies in serum using a time-resolved fluorescence immunoassay.

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is spreading rapidly around the world. Antibody detection plays an important role in the diagnosis of COVID-19. Here, we established a new time-resolved fluorescence immunoassay (TRFIA) to determine COVID-19 total antibodies. A double-antigen sandwich TRFIA was optimized and established: recombinant nucleocapsid phosphoprotein (N protein) and spike protein (S protein) of COVID-19 immobilized on 96-well plates captured human COVID-19 antibodies and then banded together with the N/S proteins labeled with europium(III) (Eu3+ ) chelates, and finally, time-resolved fluorometry was used to measure the fluorescence values. We successfully established a TRFIA method for the detection of human COVID-19 total antibodies, and the cutoff value was 2.02. There was no cross-reactivity with the negative reference of the National Reference Panel for IgM and IgG antibodies to COVID-19. The CV of the precision assay was 3.19%, and the assay could be stored stably for 15 days at 37°C. Compared with that of the colloidal gold method and chemiluminescence method, the sensitivity of the TRFIA method was higher, and the false positive/negative rate was lower. This established TRFIA has high sensitivity, accuracy, and specificity, which indicates that this method provides a new detection method for the high-throughput routine diagnosis of COVID-19.

A New Method for Detection African Swine Fever Virus: Time-resolved Fluorescence Immunoassay.

African swine fever (ASF) has severely influenced the swine industry of the whole world. Fast and accurate African swine fever virus (ASFV) antigen detection is very important for ASF prevention. This study aims to establish a new detection method for detection ASFV antigen using time-resolved fluorescence immunoassay (TRFIA) in the nose and mouth discharge. A double antibody sandwich TRFIA method was optimized and established. Recombinant P30 recombinant antigen was captured by its antibodies immobilized on 96-well plate, and then banded together with another detection antibodies labeled with Europium(III) (Eu3+) chelates, finally time-resolved analyzer measured the fluorescence intensity. The performance of this TRFIA (sensitivity, specificity and accuracy) was evaluated using the clinical samples and compared with the nucleic acid testing method. The sensitivity of this TRFIA was 0.015 ng/mL (dynamic range 0.24-500 ng/mL) with high specificity. The recovery ranged from 92.00 to 103.62 %, the inter-assay CVs ranged from 5.50 to 11.96 %, and the intra-assay CVs was between 5.20 and 10.53 %. Additionally, the cutoff value was 0.016. TRFIA took only 45 min to generate results, and its detection capability comparable to the nucleic acid detection. This study developed a TRFIA method that could be used for qualitative/quantitative detection of ASFV antigen in pigs nasal discharge, which has high sensitivity, specificity and accuracy. This TRFIA provides a new method for rapidly screening ASFV infection in pigs industry.

Time-resolved Fluorescence Immunoassay (TRFIA) for the Simultaneous Detection of MMP-9 and Lp-PLA2 in Serum.

Currently, atherosclerosis accounts for the majority of cardiovascular morbidity and mortality worldwide, and predicting the stability of atherosclerotic plaque is the main method to prevent atherosclerotic death. This study aims to establish a dual-label time-resolved fluorescence immunoassay (TRFIA) of matrix metalloprotein-9 (MMP-9) and lipoprotein-associated phospholipaseA2 (Lp-PLA2) to predict atherosclerotic plaque stability. A dual-label TRFIA was introduced for the simultaneous quantification of MMP-9 and Lp-PLA2 using fluorescent lanthanide (Eu3+ and Sm3+) chelates. The performance (sensitivity, specificity, accuracy, precision and reference intervals in different subjects) of this TRFIA was evaluated and compared with commercial kit. The sensitivity of the TRFIA for MMP-9 was 0.85 ng/mL and for Lp-PLA2 was 0.68 ng/mL with high affinity and specificity. The average recoveries were 94.58% to 109.82%, and 104.32% to 109.26%, respectively. All intra- and inter-assay CVs ranged from 3.10% to 5.46%. For the normal subjects, the cutoff value was 160.70 ng/mL for MMP-9 and 183.73 ng/mL for LP-PLA2; for the subjects with stable plaque, the cutoff value was 181.98~309.22 ng/mL for MMP-9 and 194.73~337.89 ng/mL for LP-PLA2; for the subjects with unstable plaque, the cutoff value was 330.43 ng/mL for MMP-9 and 343.23 ng/mL for LP-PLA2. This TRFIA detection results agreed well with the results of commercial kit (R2=0.9567 and R2=0.9771, respectively) in clinical serum samples. The TRFIA developed has a wide detection range and good sensitivity for the high-throughput simultaneous detection of MMP-9 and Lp-PLA2 in serum, which provides a new method for predicting the stability of atherosclerotic plaque.

Determination of parvovirus antigen in the vaccine using time-resolved fluorescence immunoassay.

As a highly contagious and potentially fatal disease of dogs, canine parvovirus type 2 (CPV-2) usually causes severe myocarditis and gastroenteritis, while vaccine injection has greatly reduced the incidence of CPV-2 diseases. However, there is currently a lack of simple and effective method for quantitative detection of CPV-2 in vaccine. Therefore, this study aims to prepare an accurate method to determine the CPV-2 antigen (CPV-2-Ag) in vaccine. Here, a sandwich time-resolved fluorescence immunoassay (TRFIA) was established and optimized. Anti-CPV-2 antibodies were immobilized on 96-well plates to capture CPV-2-Ag, and then bound together with the detection antibodies labeled with Europium(III) (Eu3+ ) chelates; finally, time-resolved fluorometry was employed to measure the fluorescence intensity. Vaccination was performed to evaluate the relationship between CPV-2-Ag concentration and antibody titer. The sensitivity is 1.15 mEU/mL (LogY = 1.524 + 0.8667 × LogX, R2  = 0.9933), and the average recovery is among 91.00% to 106.39% without cross-reactions with the other canine viral antigen. The correlation between ELISA assay and this method is up to 0.9861. And, there is high correlation between the CPV-2-Ag concentration and antibody titers (R2  = 0.9234). This immunoassay established has high sensitivity, accuracy, and specificity, which indicate that this method could be suitable for quantitative detection of CPV-2-Ag in vaccine evaluation.

HOXC10 promotes cell migration, invasion, and tumor growth in gastric carcinoma cells through upregulating proinflammatory cytokines.

HOXC10 plays a critical role in many cellular processes, such as proliferation, migration, and invasion, but the function of HOXC10 in gastric carcinoma is not clear. In this study, we aimed to investigate the expression profile of HOXC10 and its role in gastric carcinoma cells and in vivo experiments. HOXC10 expression patterns were detected in clinical samples and gastric cancer cells lines by reverse transcriptase polymerase chain reaction assays, and then, we focused on its role in regulating cell proliferation, cell cycle, migration, and invasion after transfection of silencing and overexpression plasmids in vitro and in vivo. Finally, we confirmed the correlation between HOXC10 and nuclear factor-κB (NF-κB), tumor necrosis factor-α (TNF-α), transforming growth factor-ß (TGF-ß), and epidermal growth factor receptor expression. We found that HOXC10 expression increased in clinical samples, especially in poorly differentiated (PD) gastric cancer cells. Silencing HOXC10 suppressed proliferation, migration, and invasion in vitro, and inhibited tumor growth and induced apoptosis in vivo. Overexpression of HOXC10 showed the opposite effect on PD gastric cancer cells. In addition, silencing HOXC10 inhibited the expression of interleukin-6, TNF-α, TGF-ß, and epidermal growth factor, and overexpressing HOXC10 induced their expression both in vitro and in vivo. Luciferase reporter assays and chromatin immunoprecipitation indicated that HOXC10 may activate the NF-κB signaling pathway through regulation of P65 transcriptional activity by binding to the P65 promoter. HOXC10 may play an important role in PD gastric carcinoma cell proliferation, cell cycle, migration, invasion, and metastasis through upregulating proinflammatory cytokines via NF-κB pathway, suggesting HOXC10 may serve as a novel therapeutic target for PD gastric cancer.

Determination of parvovirus antibodies in canine serum using magnetic bead-based chemiluminescence immunoassay.

Canine parvovirus type 2 (CPV-2), as a highly contagious and potentially fatal disease of dogs and many other carnivores, usually causes severe gastroenteritis and myocarditis. Therefore, it is very necessary and urgent to have an accurate method to determine the CPV-2 antibodies (CPV-2-Ab) in canine samples. Here, a magnetic bead-based chemiluminescence immunoassay was established and optimized to detect the concentration of CPV-2-Ab in serum. And a commercial assay was also used to evaluate the consistency with our method. After optimization of the detective system, the CPV-2-Ab was captured by CPV-antigen-magnetic bead (8.3 µg/mL); then combined with the conjugation of anti-canine IgG antibody-acridinium ester (0.36 µg/mL). Finally, collected the signal (read the luminosity) after 1 H reaction time. The linear correlation coefficient (R2 ) is 0.9924. The limit of detection (sensitivity) is 0.36 ng/mL (the linear dynamic range: 1.32-93.75 ng/mL), and the average recovery is 100.89% without cross-reactions with other canine viral antibodies. The results' correlation between commercial assays and this method is 0.9888. This immunoassay establishes that it has high sensitivity, accuracy, and specificity in clinical analysis, indicating that this method could be suitable for quantitative detection of CPV-2-Ab and evaluation of vaccination effect.

A dual-label time-resolved fluorescence immunoassay for the simultaneous determination of carcinoembryonic antigen and squamous cell carcinoma antigen.

Among women worldwide, cervical cancer is the second-most common cancer, and cervical smears and DNA detection have low sensitivity or are too expensive. The concentrations of carcinoembryonic antigen (CEA) and squamous cell carcinoma (SCC) in the serum were detected using a sandwich immunoassay. The CEA and SCC in the serum were captured by anti-CEA and anti-SCC antibodies. After combining other anti-CEA- and anti-SCC-labeled antibodies with europium (III) (Eu3+ ) and samarium (III) (Sm3+ ) chelates, CEA and SCC were detected with time-resolved fluorometry (TRF). The linear correlation coefficients (R2 ) of the CEA and SCC standard curves were 0.9997 and 0.9997, respectively. The minimum detection level for CEA was 1.15 ng/mL (the linear dynamic range was 3.24-543.67 ng/mL), and the average recovery was 100.83%. The sensitivity for SCC detection was 0.54 ng/mL (the linear dynamic range was 2.47-96.58 ng/mL), and the average recovery was 101.02%. High R2 between the results of commercial assays and this method were obtained (R2  = 0.9983 for CEA, R2   = 0.9878 for SCC). These findings indicated that the dual-label TRFIA invented in this study has high sensitivity, accuracy, and specificity in clinical analysis, which indicates that this method could be used for the early diagnosis and follow-up surveillance of cervical cancer.

EphB3-targeted regulation of miR-149 in the migration and invasion of human colonic carcinoma HCT116 and SW620 cells.

microRNAs play key roles during various crucial cell processes such as proliferation, migration, invasion and apoptosis. Also, microRNAs have been shown to possess oncogenic and tumor-suppressive functions in human cancers. Here, we describe the regulation and function of miR-149 in colorectal cancer cell lines. miR-149 expression patterns were detected in human colorectal cell lines and tissue samples, and then focused on its role in regulation of cell growth, migration, invasion, and its target gene identification. Furthermore, the function of the target gene of miR-149 was analyzed in vitro and in vivo. miR-149 expression was downregulated in human colorectal cancer HCT116 and SW620 cell lines compared to the normal colon epithelial NCM460 cell line using quantitative real-time polymerase chain reaction methods. Further studies indicated that introduction of miR-149 was able to suppress cell migration and invasion. Then, EphB3 was identified as a direct target gene of miR-149 in colorectal cancer cells. Moreover, experiments in vitro showed that knockdown expression of EphB3 could suppress cell proliferation and invasion, and ectopic expression of EphB3 restored the phenotypes of CRC cell lines transfected with miR149. In addition, silencing of EphB3 significantly affected cycle progression distribution and increased apoptosis in CRC cell lines. Finally, in vivo results demonstrated that knockdown of EphB3 by siRNA inhibited tumor growth. In conclusion,the important role of miR-149 in colorectal cancer progression suggesting that miR-149 may serve as a therapeutic target for colorectal cancer treatment.

The development of dual-label time-resolved fluorescence immunoassay (TRFIA) for screening of ovarian cancer based on simultaneous detection of human epididymis protein-4 and cancer antigen 125.

A two-step dual-label TRFIA was developed for the simultaneous detection of human epididymis protein-4 and cancer antigen 125 in a single run. The performance of this assay was first evaluated using clinical serum samples, and then compared with commercialized kits. The sensitivity of this assay for cancer antigen 125 detection was 0.5 U/mL (dynamic range, 0-1400 U/L), and the sensitivity for human epididymis protein-4 detection was 1 pM (dynamic range, 1-900 pM). High correlation coefficients (R) were obtained between the present dual-label TRFIA and commercially available kits (R = 0.99). The present dual-label TRFIA has high sensitivity, specificity, and accuracy in clinical sample analysis. It is a good alternative to the single-label diagnostic methods.

Development of a fluorescence immunochromatography method for quantitative measurement of matrix metalloproteinase-9.

Objective: Abnormal serum matrix metalloproteinase-9 (MMP-9) levels are closely related to the occurrence and development of many diseases. This study aimed to establish a fluorescence immunochromatography (FIC) method using the lanthanide fluorescent element europium(III) (Eu3+) for the quantitative measurement of MMP-9 in serum. Design & Methods: The FIC method for quantifying MMP-9 was optimized and established, and the FIC test strips (FICTS) were assembled and subsequently evaluated for sensitivity, specificity and precision. Furthermore, the reference interval and clinical sensitivity/specificity were estimated using clinical healthy/positive serum samples, and a commercial ELISA was used for comparison. Results: We successfully established an FIC method and prepared FICTS. The analytical sensitivity of the FICTS was 0.92 ng/mL, with a linearity range of 0-1000 ng/mL. The cross-reactivity of the 7 common serum interferents was less than 1.56%. All recoveries of the intra-array and inter-array samples ranged from 102.50% to 110.99%, and all CVs were less than 5%. The reference interval of the FICTS was >161.15 ng/mL. The clinical sensitivity was 96.00%, and the specificity was 97.5%. The results of 270 clinical serum samples were highly coincident with the clinical diagnostic results. Pearson correlation analysis and Bland‒Altman plots indicated that the FICTS and commercial ELISA results were consistent with the quantitative MMP-9 concentration. Conclusions: The designed FIC method and test strips may be suitable for point-of-care quantitative measurement of MMP-9, which provides a new method for screening for atherosclerosis, xerophthalmia, etc.

A 43.5dB Gain Unipolar a-IGZO TFT Amplifier with Parallel Bootstrap Capacitor for Bio-signals Sensing Applications.

In this paper, a high gain amplifier with phase compensation loop is presented. A structure of parallel gate cross-coupled transistors to both ends of differential pair drain and source is designed to improves the load impedance, which obtains sufficient gain and further reduces power consumption. A novel capacitor bootstrap load circuit is proposed. The capacitor bootstrap topology is constructed by the drain source resistance of the transistor working in the cut-off region, where the gate source parasitic capacitor of the transistor is in parallel with the bootstrap capacitor rather than the existing series structure, thereby only a small bootstrap capacitor is required. By avoiding the use of large capacitors, chip area can be effectively reduced without compromising performance such as gain and bandwidth. The amplifier is fabricated using 10-µm n-type a-IGZO TFT technology. Measurement results show that the proposed amplifier achieves a voltage gain of 43.5dB and a common mode rejection ratio of 61.2dB while maintaining low power consumption. The amplifier also exhibits a -3dB bandwidth covering 0.4~2.1KHz, encompassing major bioelectric frequency bands. A real-time ECG signal was successfully captured using the fabricated TFT amplifier and gel electrodes. It has great potential in flexible sensing and acquisition applications such as electro cardiogram (ECG), electro encephalogram (EEG), pulse detection, and other wearable applications.

Synergistic action of gemcitabine and celecoxib in promoting the antitumor efficacy of anti-programmed death-1 monoclonal antibody by triggering immunogenic cell death.

Background: Emerging evidence suggests that immunogenic chemotherapy not only kills tumor cells but also improves the immune-suppressive tumor microenvironment by inducing immunogenic cell death (ICD), leading to sustained anti-tumor effects. The lack of ICD inducers explored in lung cancer necessitates investigation into new inducers for this context, therefore, this study aims to explore whether the gemcitabine (GEM) and celecoxib can activate the immunogenic chemotherapy progress in lung cancer tissue. Methods: We assessed five chemotherapeutic agents for their ability to trigger ICD using ex vivo and in vivo experiments, including western blotting (WB), flow cytometry, and tumor preventive vaccine assays. Additionally, we evaluated the synergistic effects of GEM, celecoxib, and anti-programmed death 1 monoclonal antibody (aPD-1) in tumor-bearing mice to understand how GEM activates antitumor immunity and enhances immunochemotherapy. Results: GEM was identified as an effective ICD inducer, showing high expression of calreticulin (CRT) and heat shock protein 90 (HSP90). Co-culture with GEM-treated cells [Lewis lung carcinoma (LLC) and CMT-64] enhanced dendritic cell (DC) activity, evidenced by maturation markers and increased phagocytic capacity. Moreover, celecoxib was found to enhance ICD by reducing indoleamine 2,3-dioxygenase 1 (IDO1) expression and increasing reactive oxygen species (ROS)-based endoplasmic reticulum (ER) stress. The combination therapy [GEM, celecoxib, and aPD-1 (GCP)] exhibited potent and sustained antitumor activity in immunocompetent mice, with enhanced recruitment of tumor-infiltrating lymphocytes. Conclusions: These findings support the potential use of GCP therapy as a treatment option for lung cancer patients.

Senescence-related signatures predict prognosis and response to immunotherapy in colon cancer.

Background: Colorectal cancer (CRC) is one of the most common cancers. Cellular senescence plays a vital role in carcinogenesis by activating many pathways. In this study, we aimed to identify biomarkers for predicting the survival and recurrence of CRC through cellular senescence-related genes. Methods: Utilizing The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases, RNA-sequencing data and clinical information for CRC were collected. A risk model for predicting overall survival was established based on five differentially expressed genes using least absolute shrinkage and selection operator-Cox regression (LASSO-Cox regression), receiver operating characteristic (ROC), and Kaplan-Meier analyses. The study also delved into both the tumor microenvironment and the response to immunotherapy. Moreover, we gathered clinical sample data from our center in order to confirm the findings of public database analysis. Results: Through ROC and Kaplan-Meier analyses, a risk model was developed using five cellular senescence-related genes [i.e., CDKN2A, SERPINE1, SNAI1, CXCL1, and ETS2] to categorize patients into high- and low-risk groups. In the TCGA-colon adenocarcinoma (COAD) and GEO-COAD cohorts, the high-risk group was associated with a bleaker forecast (P<0.05), immune cell inactivation, and insensitivity to immunotherapy in IMvigor210 database (http://research-pub.gene.com/IMvigor210CoreBiologies/). Clinical samples were then used to confirm that ETS2 and CDKN2A could serve as independent prognostic biomarkers in CRC. Conclusions: Gene signatures related to cellular senescence, specifically involving CDKN2A and ETS2, are emerging as promising biomarkers for predicting CRC prognosis and guiding immunotherapy.

Determination of echinococcosis IgG antibodies using magnetic bead-based chemiluminescence immunoassay.

Echinococcosis is a kind of parasitic disease shared by humans and animals. The aim of this study was to establish a new detection method for echinococcosis screening using magnetic bead-based chemiluminescence immunoassay (CLIA). A magnetic bead-based CLIA to determine anti-echinococcosis IgG antibodies was optimized and established. The sensitivity, accuracy, precision and recovery rate were evaluated using the national reference serum, and the reference interval, specificity and comparison assays were performed using the clinical negative/positive echinococcosis serum samples. This study established a new CLIA to determine anti-echinococcosis IgG antibodies. The sensitivity of this CLIA method was higher than that of the registered ELISA kit and the national standard, the conformance rate of the negative/positive references was 100% (8/8), the CVs of the sensitivity reference were all below 5%, and the CVs of the precision reference were 5.7%. There was no obvious cross-reactivity with the common parasitic disease-positive serum and serum interferents. Clinical sample testing found that the cutoff value of this CLIA was 5537.15 (RLU), and there was no significant difference between the CLIA method and the registered ELISA kit. This study established a fully automated CLIA method with high sensitivity, specificity, accuracy, precision, recovery rate, and satisfactory clinical testing performance, which may provide a new choice for echinococcosis screening.

A New Method for Early Screening of Gastric Cancer (G17 and CA724 Dual-Labeled Time-Resolved Fluorescence Immunoassay).

Gastric carcinoma (GC) is one of the most common malignancies in the world with the great early screening challenges. The study is aimed at establishing a new detection method for early screening GC using time-resolved fluorescence immunoassay (TRFIA) via quantitative detection of gastrin-17 (G-17) and carbohydrate antigen 724 (CA724) in serum. Time-resolved analyzer measured the fluorescence intensity. The standards of G-17/CA724 were used for drawing the standard curve, which is used to calculate the concentration of G-17 and CA724 in serum sample. The sensitivity for G-17 was 0.54 pg/mL and for CA724 was 0.28 U/mL with a wide-range analyze concentration (0.1-1000) pg/mL or U/mL. The average recoveries ranged from 100.52% to 110.30% for G-17 and 103.02% to 116.00% for CA724. All CVs of the intra- and interassay were below 10% with high specificity. There was a high Pearson coefficient between this TRFIA method and the commercially available kits (Pearson r 0.9117 for G-17 and 0.9449 for CA724). Additionally, the cutoff value was 88.41 pg/mL and 5.47 U/mL for CA724 in health subjects. This study established a TRFIA method for simultaneous detection of the concentrations of G-17 and CA724 in serum, which provide a new method for sensitive, accurate, and specific early screening of gastric cancer.