Objective To detect anti-HCV in serum of hepatic disease patients by performing the confirmatory test, and further to confirm HCV infection. Methods Two recombinant immunoblot assays (CWT and CHIRON RIBA HCV 3.0 Strip Immunoblot Assay) were used respectively to detect anti-HCV in 477 human serum samples, which comprised 350 HCV-infected patients' specimens, 7 none-A none-E hepatitis specimens, 30 HBV-infected patients' specimens, 30 hepatitis E virus infected patients'specimens, and 60 specimens drawn from blood donors. The latter three groups served as controls. Results A total of 120 control non-HCV-infected patients' specimens were negative when tested by both assays. Among 350 HCV-infected patients, 341 were positive and 9 were indeterminated by CWT assay; 343 were positive and 7 were indeterminated by CHIRON RIBA HCV 3. 0 SIA. Seven none-A none-E hepatitis specimens tested by both assays turned out to be 2 positive, 4 negative and 1 indeterminate. The consistency rate of these two assays was 99. 16% (Kappa=0.98). Conclusion CWT assay is highly coherent with CHIRON RIBA HCV 3.0 SIA assay in the methodology of anti-HCV antibody detection, which can be applied in the determination of HCV infection among none-A none-E hepatitis patients.

Objective To express the recombinant caspid of genotype 4 hepatitis E virus(HEV) ORF2. Methods HEV recombinant capsid protein D66 was expressed in E. coli, using the ORF2 fragment (aa368-606, obtained from swine bile) of genotype 4 HEV. Results The recombinant capsid proteins D66 self-assemble to be particle with a radius of 13 nm through dimeric form in neutral solution. Coated particles reacted well with sera obtained from patients during acute or recovered phase of HEV infection. Immunofluorescence and immnoblot assay suggested that D66 bound and penetrated HepG2 cell lines, and the process of attachment was blocked by sera collected from patients during acute or recovered phase of HEV infection.Conclusion Recombinant D66 particles simulate the structure at the surface of genotype 4 HEV well and specifically adhere and penetrate the host cells, which lays the foundation for the investigation of the molecular mechanism of genotype 4 HEV infection.

Reverse genetics approaches can directly manipulate the genome of virus at the gene level, making it possible to quickly, directly and thoroughly study the mechanisms of virus replication and pathogenesis. At present, many viruses of the family Reoviridae, such as mammalian orthoreovirus ( MRV) and bluetongue virus ( BTV) , have made great progress in basic viral research using the powerful tool of re-verse genetics. However, for members of the genus Rotavirus in the family Reoviridae, progress in the con-struction of reverse genetic systems has been slow. The remarkable reverse genetics system based on helper-viruses was established in 2006, and it was not until 2017 that the entirely plasmid-based reverse genetics system was successfully established. This paper briefly reviewed the development of reverse genetics systems for rotavirus and prospected the direction for future research in order to provide technical support for acceler-ating the basic research on mechanisms of rotavirus infection.

We developed a high-sensitivity C-reactive protein quantifiable chemiluminescent immunoassay (hs-CRP CLIA). The high-purity native CRP was purified from hepatic cirrhosis patient ascetic fluid by affinity and ion exchange chromatography and used as an immunogen to develop the monoclonal antibodies (mAbs) against CRP. Twenty-two mAbs were identified reactive with CRP in ELISA and 13 of them were reactive in the phosphorycholine ligand capture ELISA. The mAbs 10C5 and 10C11 were selected to develop the hs-CRP CLIA. The linearity and performance of the hs-CRP CLIA was characterized. It was showed not reactive when testing against other serum materials (IgG, hemoglobin and triglyceride). The reliable correlation (R2 > 0.993) was obtained between testing value (RLU/S) and the concentration of human serum CRP calibrator. The linearity fell in the range of 0.04-20.38 mg/L. The assay has good accuracy and reproducibility, the mean recovery was 99% and the precision of the intra- and inter assay was CVs (4.2%-5.8%) and (9.0%-11.5%), respectively. In testing of 90 human sera, this assay performed well and correlated comparably with a commercial hs-CRP ELISA kit. Thus, hs-CRP CLIA is an accurate, reliable, quantifiable assay for detection of high-sensitive C-reactive protein in serum, it may be useful to improve the risk assessment of cardiovascular disease and the prognosis of inflammatory bowel disease.
C-Reactive Protein ; analysis ; chemistry ; Humans ; Immunoassay ; methods ; Luminescent Measurements ; methods ; Sensitivity and Specificity

Objective:To understand the pathogenic situation of hand, foot and mouth disease (HFMD) in Linyi City, and to analyze the gene characteristics of Coxsackievirus A10 complete VP1 region.Methods:The samples of HFMD cases from Linyi City in 2023 were tested for typing and strain isolation, and the VP1 gene of CV-A10 isolate was amplified and sequenced. The obtained sequences were compared with those in the NCBI database, and the phylogenetic tree was constructed for gene characteristics and molecular epidemiological analysis.Results:In 2023, a total of 861 samples of HFMD were collected, and 594 (68.99%) were positive for nucleic acid tests. The male to female ratio of positive cases was 1.56∶1. Children under 5 years old accounted for 81.65%, and the highest incidence season was from June to August (83.84%). CV-A6 was the main pathogen (84.51%), followed by CV-A10 (9.93%). The nucleotide and amino acid homology of VP1 gene sequence among 13 isolates were 93.29%-100.00% and 97.65%-100.00%, respectively. The nucleotide and amino acid homology with AF081300-Kowalik/USA/1950 was lower (75.95%-76.62%, 91.72%-92.41%). The amino acid and nucleotide homology with C2c was the highest (94.28%-96.76%, 98.28%-100.00%), and the genetic distance was the closest (0.04-0.06). Amino acid site variation analysis showed that compared with the prototype strain AF081300-Kowalik/USA/1950, the isolates had more site variation, while only some isolates had I80V, E141K, P147S, T219I, E240K and V261I mutations compared with the representative strain C2c. The genetic evolution tree showed that the isolates were all in the same clade as the C2c representative strains, and they all belonged to the C2c genotype, and the isolates were further divided into two smaller clades.Conclusions:In 2023, CV-A6 was the main pathogen of HFMD in Linyi City, followed by CV-A10. All CV-A10 isolates were C2c genotypes and can be divided into two evolutionary clades. Continuous monitoring and genetic characterization of CV-A10 should be strengthened.

With the advantages of low immunogenicity and long half-life, human monoclonal antibody has become an indispensable biological agent in vivo. Immortalization of human B cells is a potential and effective method to obtain natural human antibody library, which can provide a rich source for the preparation of human monoclonal antibodies. As there are urgent problems to be solved in each platform, the preparation of antibodies based on human B cell immortalization is still limited to the laboratory research stage. At present, there is a lack of a systematic review to clarify the advantages and disadvantages of the existing human B cell immortalization antibody preparation platform and its feasibility analysis. This paper reviews the research on the preparation of human monoclonal antibody based on human B cells immortalization, and describes an in vitro cell culture method, in which hCD40L vesicles are used instead of feeder cells, in order to provide references for the further development of human monoclonal antibody preparation technology.
Antibodies, Monoclonal ; B-Lymphocytes ; Cell Culture Techniques ; Humans

In vitro diagnosis (IVD) is an important source of clinical diagnostic information, and provides an important decision basis for disease prevention, diagnosis and treatment. IVD is a necessary tool for promoting graded diagnosis and treatment, realizing precision medicine, constructing a "Healthy China" and responding to major public health emergencies. Combining the great progress made in the development of in vitro diagnostics in China and the shortcomings and weaknesses faced by it, this article analyzed the demand for IVD, policy support, technical and industrial development trends, and the ways to accelerate the industrialization development, aiming to promote the development and improvement of IVD in China.

Rotavirus (RV) is one of the leading causes of acute gastroenteritis in infants and young animals worldwide. Rotavirus infection has obvious species specificity and mainly causes diarrhea in infants and young animals. The host innate responses suppress the infection and replication of rotavirus through activating multiple signaling pathways. Meanwhile, rotavirus also antagonizes the innate immune responses in various ways. This article reviewed the mechanisms of host innate immune responses to rotavirus infection and the antagonistic mechanism of rotavirus against host innate immunity with a view to providing reference for the development of therapeutic drugs and the prevention of rotavirus infection.

With the requirements of early diagnosis, biomarker development and functional definition, the challenge of sensitivity of immunoassay has become increasingly prominent. How to improve it to break the bottleneck has become a major challenge in the field of bioassays. Amplifying the immunosignal is the most direct method to improve detection sensitivity. Biotin-avidin system (BAS), tyramide signal amplification (TSA) and immuno-polymerase chain reaction (Im-PCR) are the most classic signal amplification techniques which significantly improved the sensitivity of immunoassays. In recent years, studies have confirmed that the sensitivity of immunoassays can be further increased by approximately three orders of magnitude with the invention of techniques including catalyzed reporter deposition-based signal amplification, nanotechnologies-based signal amplification and hybridization chain reaction-based signal amplification. Herein, we will summarize the techniques that have been developed in recent years for amplifying the signals of immunodetection and comparatively analyze their advantages and disadvantages in order to provide reference for the developed techniques transformed to clinical application and further research on ultrasensitive immunoassays.

Rotavirus is one of the most common causes of gastroenteritis among children under five years of age worldwide. Most children are infected with rotavirus in their early childhood. Currently, no ef-fective treatment is available for diarrhea caused by rotavirus infection, and vaccination is the most effective way to prevent rotavirus infection. With the introduction of rotavirus vaccines, the morbidity and mortality of severe diarrhea in children caused by rotavirus infection are declined significantly, but these rotavirus vac-cines have significantly lower efficacy in developing countries in Africa and some countries in Asia where the mortality of rotavirus-related diseases is high than in developed countries. High titers of maternal antibodies may be one of the reasons why the efficacy of rotavirus vaccines is low in these countries. Extensive studies have been conducted on the relationship between neonatal response to rotavirus vaccine immunization and ro-tavirus-specific antibodies in the mother′s milk and serum in recent years. However, results of some studies are in conflict. In this review, we summarize the relationship between maternal antibodies and the immune responses after vaccination with rotavirus vaccines in order to provide basis for improving the efficacy of rota-virus vaccines in low-income countries.