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Immunofluorescence Lateral Chromatography in Combination

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Immunofluorescence Lateral Chromatography in Combination Materials Express 2158-5849/2021/11/248/007 Copyright © 2021 by American Scientific Publishers All rights reserved. doi:10.1166/mex.2021.1886 Printed in the United States of America www.aspbs.com/mex Immunofluorescence lateral chromatography in combination with detection of hypersensitive C-reactive protein and procalcitonin levels in patients with mental disease associated with coronavirus 2019 disease infection Dan Li1,†, Xiaoliang Zhou2,†,YunShi1,†,PingLi1, Shuzi Chen1, Weifeng Jin1, Xiaoyan Lou1, Zhenhua Li1, Ping Lin1,∗, and Yang Sun3,4,∗ 1Department of Clinical Laboratory, Shanghai Mental Health Center, Shanghai 200030, PR China 2Department of Clinical Laboratory, Wuhan Mental Health Center, Wuhan 430022, Hubei, PR China 3Institute of Arthritis Research, Shanghai Academy of Chinese Medical Sciences, Shanghai 200052, PR China 4Guanghua Integrative Medicine Hospital, Shanghai 200052, PR China IP: 192.168.39.210 On: Sun, 26 Sep 2021 07:58:46 Copyright: American Scientific Publishers Delivered by Ingenta ABSTRACT Article Immunofluorescence lateral chromatography (IFLC) is a quantitative detection technique used to screen C-reactive protein and procalcitonin (PCT) monoclonal antibodies with high efficiency and specificity. This study investigated the application value of IFLC combined with the detection of hypersensitive C-reactive protein (hsCRP) and PCT in patients with mental diseases associated with COVID-19 infection. Sixty-four patients with mental disease admitted to our hospital from January to April 2020, were selected as research subjects. Based on the diagnostic criteria for COVID-19 in the “Corona Virus Disease 2019 Diagnosis and Treatment Scheme (Byass, P., 2020. Eco-epidemiological assessment of the COVID-19 epidemic in China, January- February 2020. Global Health Action, 13(1), p.1760490.) version 6, all patients were tested using the nucleic acid amplification test for COVID-19 and were divided into two groups of 6 infected and 58 non-infected patients. Subsequently, the patients’ hsCRP and PCT levels were measured using IFLC and an automatic biochemistry analyzer (BA). The results showed that the hsCRP and PCT levels in the infected group were higher than the non-infected group. Additionally, the analytical ultracentrifugation (AUC), sensitivity, and specificity of hsCRP were higher than PCT. The AUC, sensitivity, and specificity of hsCRP+PCT measured jointly were significantly higher than hsCRP and PCT measured separately. The BA results showed that the AUC, sensitivity, and speci- ficity of hsCRP were higher than PCT. The AUC, sensitivity, and specificity of hsCRP + PCT measured jointly were significantly higher than hsCRP and PCT measured separately. Comprehensive analysis showed that the AUC, sensitivity, and specificity of hsCRP+PCT tested using IFLC were significantly higher than using the BA. All of the above differences were statistically significant (P<0.05). It was concluded that the joint detection of hsCRP and PCT by IFLC helped identify and diagnose mental diseases associated with COVID-19 infection. ∗Authors to whom correspondence should be addressed. †These three authors contributed equally to this work. 248 Mater. Express, Vol. 11, No. 2, 2021 Immunofluorescence lateral chromatography Materials Express Li et al. Keywords: Immunofluorescence, Joint Detection, COVID-19, Hypersensitive C-Reactive Protein, Procalcitonin. 1. INTRODUCTION 2. MATERIALS AND METHODS When the blood or respiratory tract samples tested using 2.1. Materials real-time fluorescence quantitative polymerase chain reac- 2.1.1. Patient Information tion are positive for the novel Coronavirus 2019, or The patients with mental diseases admitted to the Wuhan their viral gene sequencing is significantly homologous Mental Health Center, Wuhan, China, from January to to it, patients are confirmed to be infected with COVID- April 2020, were selected as the research subjects. Inclu- 19 (Coronavirus 2019 Disease) [1]. Mental illness is a sion criteria: (1) meet the diagnostic criteria for mental ill- special kind of disease. A patients’ mental state may ness [5]; (2) the age of 18 years old or more; (3) no recent be affected by their perception of COVID-19 symptoms history of surgery or trauma; (4) family members sign that could even lead to the aggravation of hypochondria, informed consent forms. Exclusion criteria: (1) concomi- social phobia, generalized anxiety disorder, and obsessive- tant immune system diseases; (2) combined with infec- compulsive disorder, which is not conducive to the treat- tious diseases other than COVID-19; (3) complicated with ment of mental illness [2]. COVID-19 is characterized malignant tumor; (4) suffering from heart, liver, kidney by fever, dry cough, fatigue, and other clinical manifesta- and other important visceral lesions. Sixty-four patients tions. It is highly contagious and insidious and can occur were enrolled in the study. Based on the diagnostic criteria in people of all ages. Currently, the nucleic acid test is for COVID-19 in the “Corona Virus Disease 2019 Diag- frequently used in the clinical diagnosis of COVID-19, nosis and Treatment Scheme” version 6, all patients were but it is significantly affected by external factors, lead- tested using the nucleic acid amplification test [6]. They ing to some false-negative results, and there are defi- were divided into an infected group (n = 6) and a non- ciencies in sensitivity and specificity [3]. Therefore, it is infected group (n = 58). 2 detect was used to compare the Article important to find an effective, rapid, and accurate method genders of the two groups, while t test was used to com- for identifying COVID-19 infected patients so that they IP: 192.168.39.210 On: Sun,pare 26 theSep age, 2021 height 07:58:46 and body mass index, and the differ- may obtain timely and effective treatmentCopyright: and stopAmerican the Scientific Publishers ences were not statistically significant (P>0.05) (Table I). spread of the virus. The quantitative detection techniqueDelivered by Ingenta This study was approved by the Medical Ethics Committee of immunofluorescence lateral chromatography (IFLC) is of our hospital. based on the reaction principle of the double-antibody sandwich system. The technique selects monoclonal anti- 2.1.2. Reagents and Suppliers bodies against highly specific target proteins to pair with each other and then produces fluorescence-labeled conju- All reagents were used in this study were analytically · gation; the cloaked, captured antibodies form a double- pure: Na2HPO4 12H2O (disodium hydrogen phosphate) antibody sandwich system, and finally, it detects the target (article No.: 0404, Shanghai Liansuo Biotechnology Co., proteins in serum sample [4]. HsCRP is an acute phase Ltd.), triton-x100 (article No.: X-100, Haian Petrochemi- protein synthesized by liver cells; it can increase sharply cal Plant, Jiangsu Province, China), tris (article No.: 0161 when the body is infected and returns to normal range on AMRESCO), tween-20 (article No.: P137, Shanghai Lian- the remission of the disease and recovery of tissues and suo Biological Technology Co., Ltd.), 99% PROCLIN- structural functions. Therefore, the level of hsCRP reflects 300 (article No.: ZB116, Shanghai Jinjing Biotechnology the occurrence, development, and prognosis of infectious Co., Ltd.), 99% trehalose (article No.: YY11150, Shanghai diseases. PCT is a protein that increases with protozoan, Yuanye Biotechnology Co., Ltd.) polyethylene glycol 600 systemic bacterial or other bacterial infections. Therefore, (article No.: 167161, Xibao Biological Technology, Co., the occurrence and development of infectious diseases can Ltd., Shanghai), fluorescent microspheres F1-XC005 and be accompanied by in vivo changes in serum hsCRP and F1-XC03 (Hongrong Weizre Bioengineering Technology PCT levels. In this study, patients with mental illness and Co., Ltd., Shanghai), polyester fiber film, cellulose nitrate infected with COVID-19 were the research subjects, while film CN95, and glass fiber film 8964 (Xuxingtai Quan- non-infected patients with mental illness were the controls. tity Technology Co., Ltd.) PVP K30 (polyvinylpyrroli- In addition to IFLC, an automatic biochemistry analyzer done) (article No.: J0029-1, Suzhou Yaco Technology Co., (BA), commonly used in clinical practice, was used for Ltd.) and PVA (polyvinyl alcohol), article No.: S25454 testing study participants. The purpose of the study was (Shanghai Yuanye Biotechnology Co., Ltd.), CRP mon- to explore the effect of IFLC combined with hsCRP and oclonal antibodies C2, C6, PCT monoclonal antibodies PCT detection on the diagnosis of COVID-19 and provide 42, and 16B5 (Shanghai Yibo Biotechnology Co., Ltd.), a reference for the clinical optimization of the diagnostic CM-EUs (Thermo Fisher Scientific Inc.), sample pads and process. absorbent pads (Bedford, Massachusetts, USA). Mater. Express, Vol. 11, pp. 248–254, 2021 249 Materials Express Immunofluorescence lateral chromatography Li et al. Table I. Comparison of data between the infected and non-infected groups. Gender [case, (n)%] Body mass index Group Male Female Age (x ± s, year) Height (x ± s,cm) (x ± s, kg/m2) Infected group (n = 6) 4 (66.67) 2 (33.33) 45.29 ± 6.54 163.03 ± 5.02 23.05 ± 3.31 Aon-infected group (n = 58) 28 (48.28) 30 (51.72) 45.37 ± 6.47 162.89 ± 5.04 22.94 ± 3.58 2/t 0.184 0.029 0.065 0.072 P 0.668 0.977 0.949 0.943 2.2. Methods microsphere [7, 8]. After activating the fluorescent micro- 2.2.1. Quantitative Immunofluorescence Lateral spheres, the coupled buffer (0.02 M PB, pH 7.4) was taken Chromatography to wash the activated fluorescent microspheres twice ultra- The flowchart for the method is shown in Figure 1. The sonically. Then, the CRP antibody was added as required hsCRP and PCT levels of all patients were determined by to obtain the fluorescence-labeled conjugation (concentra- IFLP. The steps are as follows: the sample diluent was tion: 0.2–1 mg/mL), and the reaction container was oscil- prepared (PEG 6000: triton-X100: proclin-300: buffer = lated at 37 C for 120–180 min. The prepared coupling 0.5%:0.5%:0.02%:0.05 M PBS (Phosphate Buffer Saline), buffer was used to change the liquid and then oscillated pH 7.4), sodium phosphate buffer (0.01 M PBS, pH 7.4), continuously at 37 C for 30 min.
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