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BackgroundMore than ten novel COVID-19 vaccines have been approved with protections against SARS-CoV-2 infections ranges between 52-95%. It is of great interest to the vaccinees who have received the COVID-19 vaccines, vaccine developers and authorities to identify the non-responders in a timely manner so intervention can take place by either giving additional boosts of the same vaccine or switching to a different vaccine to improve the protection against the SARS-CoV-2 infections. A robust correlation was seen between binding antibody titer and efficacy (p=0.93) in the clinic studies of 7 COVID-19 vaccines, so it is of urgency to develop a simple POCT for vaccinees to self-assess their immune response at home. MethodsUsing CHO cell-expressed full length SARS-CoV2 S1 protein as coating antigen on colloidal gold particles, a SARS-CoV-2 S1 IgG-IgM antibody lateral flow test kit (POCT) was developed. The test was validated with negative human sera collected prior to the COVID-19 outbreaks, and blood samples from human subjects prior, during, and post-immunization of COVID-19 vaccines. ResultsThe specificity of the POCT was 99.0%, as examined against 947 normal human sera and 20 whole blood samples collected pre-immunization. The limit of detection was 50 IU/mL of pseudovirus neutralizing titer (PVNT) using human anti-SARS-2 neutralizing standards from convalescent sera. The sensitivity of POCT for SARS-CoV-2 S1 protein antibody IgG-IgM was compared with SARS-CoV-2 RBD antibody ELISA and determined to be 100% using 23 blood samples from vaccinated human subjects and 10 samples from non-vaccinated ones. Whole blood samples were collected from 119 human subjects (ages between 22-61 years) prior to, during, and post-vaccination of five different COVID-19 vaccines. Among them, 115 people tested positive for SARS-CoV-2 S1 antibodies (showing positive at least once) and 4 people tested negative (tested negative at least twice on different days), demonstrating 96.64% of seroconversion after full-vaccination. 92.3% (36/39) of the human subjects who were younger than 45 achieved seroconversion within 2 weeks while only 57.1% (4/7) of subjects older than 45 tested positive for S1 antibodies, suggesting that younger people develop protection much faster than older ones. Even though the S1 antibody level in 88% of human subjects vaccinated with inactivated virus dropped below 50 IU/mL two months later, one boost could quickly raise the S1 antibody titer above 50 IU/mL of PVNT, indicates that the initial vaccination was successful and immunization memory was developed. ConclusionUsing the lateral flow tests of SARS-CoV2 S1 IgG+IgM, vaccinated human subjects can easily self-assess the efficacy of their vaccination at home. The vaccine developer could quickly identify those non-responders and give them an additional boost to improve the efficacy of their vaccines. Vaccinees who failed in response could switch to different types of COVID-19 vaccines since there are more than 10 COVID-19 vaccines approved using three different platform technologies. HighlightsO_LIMore than ten novel COVID-19 vaccines have been approved with protections against SARS-CoV-2 infections ranges between 52-95%. It is of great interest to the vaccinees who have received the COVID-19 vaccines, vaccine developers and authorities to identify the non-responders in a timely manner. C_LIO_LIA highly specific and very simple lateral flow test kit for measurement of SARS-CoV-2 S1IgG+IgM antibodies post-immunization of COVID-19 vaccine using peripheral blood was developed as a home-test assay with a limit of detection (LOD) at 50 IU/mL of pseudovirus neutralizing titer (PVNT). C_LIO_LIAfter full vaccinations with COVID-19 vaccines, 96.6% of the volunteers successfully achieved the seroconversion of SARS-CoV-2 S1 IgG+IgM antibody. C_LIO_LI92.3% (36/39) of the human subjects who were younger than 45 achieved seroconversion within 2 weeks while only 57.1% (4/7) of subjects older than 45 tested positive for S1 antibodies, suggesting that younger people develop protection much faster than older ones. C_LIO_LIEven though the S1 antibody level in 88% of human subjects vaccinated with inactivated virus dropped below the detection 2-6 months later, one boost could quickly raise the S1 antibody titer above 50 IU/mL of PVNT, indicating that the initial vaccination was successful and immunization memory was developed. C_LI
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The COVID-19 outbreak has become a global pandemic responsible for over 2,000,000 confirmed cases and over 126,000 deaths worldwide. In this study, we examined the immunogenicity of CHO-expressed recombinant SARS-CoV-2 S1-Fc fusion protein in mice, rabbits, and monkeys as a potential candidate for a COVID-19 vaccine. We demonstrate that the S1-Fc fusion protein is extremely immunogenic, as evidenced by strong antibody titers observed by day 7. Strong virus neutralizing activity was observed on day 14 in rabbits immunized with the S1-Fc fusion protein using a pseudovirus neutralization assay. Most importantly, in less than 20 days and three injections of the S1-Fc fusion protein, two monkeys developed higher virus neutralizing titers than a recovered COVID-19 patient in a live SARS-CoV-2 infection assay. Our data strongly suggests that the CHO-expressed SARS-CoV-2 S1-Fc recombinant protein could be a strong candidate for vaccine development against COVID-19. HighlightsO_LICHO-expressed S1-Fc protein is very immunogenic in various animals and can rapidly induce strong antibody production C_LIO_LIS1-Fc protein solicits strong neutralizing activities against live virus C_LIO_LIStable CHO cell line expressing 50 mg/L of S1-Fc and a 3,000 L Bioreactor can produce 3 million doses of human COVID-19 vaccine every 10 days, making it an accessible and affordable option for worldwide vaccination C_LI
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WHO has declared COVID-19 a pandemic with more than 300,000 confirmed cases and more than 14,000 deaths. There is urgent need for accurate and rapid diagnostic kits. Here we report the development and validation of a COVID-19/SARS-CoV-2 S1 serology ELISA kit for the detection of total anti-virus antibody (IgG+IgM) titers in sera from either the general population or patients suspected to be infected. For indirect ELISA, CHO-expressed recombinant full length SARS-CoV-2-S1 protein with 6* His tag was used as the coating antigen to capture the SARS-CoV-2-S1 antibodies specifically. The specificity of the ELISA kit was determined to be 97.5%, as examined against total 412 normal human sera including 257 samples collected prior to the outbreak and 155 collected during the outbreak. The sensitivity of the ELISA kit was determined to be 97.5% by testing against 69 samples from hospitalized and/or recovered COVID-19 patients. The overall accuracy rate reached 97.3%. Most importantly, in one case study, the ELISA test kit was able to identify an infected person who had previously been quarantined for 14 days after coming into contact with a confirmed COVID-19 patient, and discharged after testing negative twice by nucleic acid test. With the assays developed here, we can screen millions of medical staffs in the hospitals and people in residential complex, schools, public transportations, and business parks in the epidemic centers of the outbreaks to fish out the "innocent viral spreaders", and help to stop the further spreading of the virus.
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The COVID-19 China coronavirus started in Dec 2019 was challenged by the lack of accurate serological diagnostic tool for this deadly disease to quickly identify and isolate the infected patients. The generation of COVID-19-specific antibodies is essential for such tasks. Here we report that polyclonal and monoclonal antibodies were generated by immunizing animals with synthetic peptides corresponding to different areas of Nucleoprotein (N) of COVID-19. The specificities of the COVID-19 antibodies were assessed by Western Blot analysis against NPs from COVID-19, MERS and SARS. Antibodies were used for immunohistochemistry staining of the tissue sections from COVID-19 infected patient, as a potential diagnostic tool. A Sandwich ELISA kit was quickly assembled for quantitation of the virus/NP of COVID-19 concentrations in the vaccine preparations. Development of POCT is also aggressively undergoing.
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Objective To study the effects of severely burned rats serum on migration of BMSCs and mechanism.Methods Severely burned rats model was established,and the preparation of severely burned rats serum.Experimental groups:normal train-ing group(containing 10% fetal bovine serum,group C),burn serum group(containing 10% burns in the rat serum,group B),burn serum+blockers(10% burns in the rat serum+final concentration of 10 μmol/L PI3K signaling pathway inhibitor LY294002 train-ing,group B+LY).Activity of cells was examined with MTT;migration of cells was examined with Transwell chambers testing;protein expression of p-AKT/AKT was determined with Western blot;microtubule structure of cells was examined with immuno-fluorescence.Results Compared with group C,group B burn serum treatment after 24 h,BMSCs activity(P <0.01),p-AKT levels (P <0.05),increased migration quantity(P <0.001);cell microtubule structures appear rupture,after adding inhibitor,compared with group B,group B+LY BMSCs activity(P <0.01),to reduce the number of migration(P <0.001),p-lower AKT(P <0.05), cell microtubule structure similar to the normal group.Conclusion Severely burned rats serum can promote BMSCs migration,may burn serum cytokine activation of PI3K/AKT signal pathway,resulting in cell microtubule structure change,promote the migration of BMSCs.
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Objective To get the experience of surgical treatment for diabetic foot (DF) ulcer.Methods Clinical data of 85 patiems (108 limbs in total) admitted in Department of Bum and Plastic Surgery in The First Affiliated Hospital of Chongqing Medical University from Jan 2010 to Dec 2012 were retrospectively analyzed.Results Wound culture results of the 85 patients were:bacteria in 142 limbs,and fungi in 7 limbs.The main bacteria cultured were staphylococcus aureus,escherichia coli,coagulase negative staphylococcus,enterococcus,pseudomonas aeruginosa,and acinetobacter baumannii.DF healing and Wagner classification was negatively correlated:the higher the class,the less the primary healing rate.The higher amputation rate were found in the higher class patients.The more delayed healing,and the longer the healing time and average hospital stay were also found in the higher class patients.85 patients underwent surgery:skin grafting in 45 cases (52.9％),flap in 8 cases (9.4％),skin repair + flap grafting in 7 cases,toe (limb) amputation in 28 cases,2 cases were sutured after debridement.All cases were cured after one or multiple surgeries.No recurrence was found during the follow-up of 10 days to 2 months.No delayed infection occurred to the repairments.The shape and weight bearing walking function were good.Conclusions The treatment of DF need to follow the principle of multidisciplinary cooperation,professional treatmem,systemic and local treatment.We need to pay attention to the etiology and prevention of the disease,focus on wound bed preparation,choose the best treatment.Early operation can significandy fascilitate wound healing.
