Porcine deltacoronavirus(PDCoV)is one of the important pathogens associated with swine viral diarrhea,which results in enormous economic losses in the pig industry.Among the known encoding proteins,the nucleocapsid(N)protein is relatively conserved and serves as the dominant antigen of coronaviruses,making it an ideal candidate for the early and accurate diagnosis of infection.In this study,the PDCoV N protein was induced and expressed in a prokaryotic system using IPTG treatment at low temperatures.Six-week-old BALB/c mice were immunized with the recombinant PDCoV N protein purified by nickel column.Isolated spleen cells were harvested and fused with SP2/0 cells once the serum titers reached 1∶10 000,as determined by indirect ELISA.The positive monoclonal hybridoma with the highest titer of secretory antibodies were screened af-ter three rounds of limited dilution.Ultimately,indirect immunofluorescent assay(IFA)and West-ern blot were employed to confirm the characterization of the antibodies,and the subtypes of the heavy and light chains of the antibodies were determined.One strain of hybridoma against the PD-CoV N protein,designated as 2G12,was obtained,with a titer approaching 1∶40 960.The IFA as-say showed PDCoV was specifically bound to 2G12.Furthermore,the Western blot assay further showed PDCoV in the supernatant of infected cells or recombinant PDCoV N protein,reacted well with 2G12.The subtype of MAbs was determined as IgG1 with the light chains being κ.The MAbs generated against the PDCoV N protein in this study provide valuable support for the development of a diagnostic kit for PDCoV and play an important role in the functional research of the PDCoV N protein.

Objective:To explore the application value of laparoscopic contrast enhanced ultrasound in laparoscopic surgery for patients with hepatocellular carcinoma combined with cirrhosis.Methods:The clinical data of 71 patients with hepatocellular carcinoma combined cirrhosis from February 2018 to February 2020 in Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology were retrospectively analyzed. The patients underwent preoperative enhanced CT and multi-parameter MRI examination, followed by laparoscopic partial hepatectomy, and intraoperative laparoscopic contrast enhanced ultrasound examination. Based on histological examination and follow-up results, the diagnostic efficacy of preoperative imaging and preoperative imaging combined with intraoperative laparoscopic contrast enhanced ultrasound in patients with hepatocellular carcinoma combined with cirrhosis was compared.Results:Among the 71 patients, 69 completed laparoscopic surgery and 2 converted to open surgery. One hundred and ten HCC lesions were diagnosed by preoperative imaging examination, 105 lesions were detected by intraoperative ultrasound among them, of which 98 lesions were diagnosed as HCC by intraoperative laparoscopic contrast enhanced ultrasound. There were no statistically significant difference in sensitivity, specificity, accuracy, positive predictive value and negative predictive value between preoperative imaging and preoperative imaging combined with intraoperative laparoscopic contrast enhanced ultrasound in the diagnosis of malignant liver lesions: 94.4% (102/108) vs. 99.1% (107/108), 81.0% (34/42) vs. 66.7% (28/42), 90.6% (136/150) vs. 90.0% (135/150), 92.7% (102/110) vs. 88.4% (107/121) and 85.0% (34/40) vs. 96.6% (28/29), P>0.05. Laparoscopic contrast enhanced ultrasound revealed an additional 11 suspected malignant lesions, of which 5 lesions were histologically confirmed as HCC. Seven patients underwent surgical strategy changes. Conclusions:Laparoscopic contrast enhanced ultrasound in patients with HCC combined with cirrhosis during laparoscopic surgery can be used to detect, identify, accurately locate of the lesions and modify the surgical plan.

Porcine deltacoronavirus(PDCoV)is one of the important pathogens associated with swine viral diarrhea,which results in enormous economic losses in the pig industry.Among the known encoding proteins,the nucleocapsid(N)protein is relatively conserved and serves as the dominant antigen of coronaviruses,making it an ideal candidate for the early and accurate diagnosis of infection.In this study,the PDCoV N protein was induced and expressed in a prokaryotic system using IPTG treatment at low temperatures.Six-week-old BALB/c mice were immunized with the recombinant PDCoV N protein purified by nickel column.Isolated spleen cells were harvested and fused with SP2/0 cells once the serum titers reached 1∶10 000,as determined by indirect ELISA.The positive monoclonal hybridoma with the highest titer of secretory antibodies were screened af-ter three rounds of limited dilution.Ultimately,indirect immunofluorescent assay(IFA)and West-ern blot were employed to confirm the characterization of the antibodies,and the subtypes of the heavy and light chains of the antibodies were determined.One strain of hybridoma against the PD-CoV N protein,designated as 2G12,was obtained,with a titer approaching 1∶40 960.The IFA as-say showed PDCoV was specifically bound to 2G12.Furthermore,the Western blot assay further showed PDCoV in the supernatant of infected cells or recombinant PDCoV N protein,reacted well with 2G12.The subtype of MAbs was determined as IgG1 with the light chains being κ.The MAbs generated against the PDCoV N protein in this study provide valuable support for the development of a diagnostic kit for PDCoV and play an important role in the functional research of the PDCoV N protein.