Mice Immunized with the Vaccine Candidate HexaPro Spike Produce Neutralizing Antibodies against SARS-CoV-2.

Updated and revised versions of COVID-19 vaccines are vital due to genetic variations of the SARS-CoV-2 spike antigen. Furthermore, vaccines that are safe, cost-effective, and logistic-friendly are critically needed for global equity, especially for middle- to low-income countries. Recombinant protein-based subunit vaccines against SARS-CoV-2 have been reported using the receptor-binding domain (RBD) and the prefusion spike trimers (S-2P). Recently, a new version of prefusion spike trimers, named HexaPro, has been shown to possess two RBD in the "up" conformation, due to its physical property, as opposed to just one exposed RBD found in S-2P. Importantly, this HexaPro spike antigen is more stable than S-2P, raising its feasibility for global logistics and supply chain. Here, we report that the spike protein HexaPro offers a promising candidate for the SARS-CoV-2 vaccine. Mice immunized by the recombinant HexaPro adjuvanted with aluminum hydroxide using a prime-boost regimen produced high-titer neutralizing antibodies for up to 56 days after initial immunization against live SARS-CoV-2 infection. Also, the level of neutralization activity is comparable to that of convalescence sera. Our results indicate that the HexaPro subunit vaccine confers neutralization activity in sera collected from mice receiving the prime-boost regimen.

Systematic comparison of four point-of-care methods versus the reference laboratory measurement of hemoglobin in the surgical ICU setting: a cross-sectional method comparison study.

BACKGROUND: Transfusion decision during the perioperative period mostly relies on the point-of-care testing for Hb measurement. This study aimed systematically compared four point-of-care methods with the central laboratory measurement of hemoglobin (LHb) regarding the accuracy, precision, and assay practicality to identify the preferred point-of-care method during the perioperative period. METHODS: This cross-sectional method comparison study was conducted in the surgical intensive care unit at Ramathibodi Hospital, Thailand, from September 2015 to July 2016. Four point-of-care methods, i.e., capillary hematocrit (HctCap), HemoCue Hb201+, iSTAT with CG8+ cartridge, and SpHb from Radical-7 pulse co-oximeter were carried out when LHb was ordered. Pearson correlation and Bland-Altman analyses were performed to assess the accuracy and precision, while the workload, turnaround time, and the unit cost were evaluated for the method practicality. RESULTS: Thirty-five patients were enrolled, corresponding to 48 blood specimens for analyses, resulting in the measured hemoglobin of 11.2 ± 1.9 g/dL by LHb. Ranking by correlation (r), mean difference (bias) and 95% limit of agreement (LOA) showed the point-of-care methods from the greater to the less performance as followed, iSTAT-LHb pair (r = 0.941; bias 0.15 (95% LOA; - 1.41, 1.12) g/dL), HemoCue-LHb pair (r = 0.922; bias - 0.18 (95% LOA; - 1.63, 1.28) g/dL), SpHb-LHb pair (r = 0.670; bias 0.13 (95% LOA; - 3.12, 3.39) g/dL) and HctCap-LHb pair (r = 0.905; bias 0.46 (95% LOA; - 1.16, 2.08) g/dL). Considering the practicality, all point-of-care methods had less workload and turnaround time than LHb, but only HemoCue and HctCap had lower unit cost. CONCLUSION: This study identified HemoCue as the suitable point-of-care method for the sole purpose of Hb measurement in the surgical ICU setting, while iSTAT should be considered when additional data is needed.

Systematic comparisons of various spectrophotometric and colorimetric methods to measure concentrations of protein, peptide and amino acid: detectable limits, linear dynamic ranges, interferences, practicality and unit costs.

There is limited and inconclusive information regarding detectable limits and linear dynamic ranges of various quantitative protein assays. We thus performed systematic comparisons of seven commonly used methods, including direct spectrophotometric quantitation at λ205 and λ280 nm (A205 and A280, respectively), bicinchoninic acid (BCA), Biuret, Bradford, Lowry and Ninhydrin methods. Purified BSA, porcine kidney extract, tryptic digested peptides derived from purified BSA, and glycine, were used as representative purified protein, complex protein mixture, peptide and amino acid, respectively. Bradford method was the most sensitive assay (LOD=0.006 mg/ml) and had the widest range of detectability (LOD-UOD=0.006-100mg/ml) for purified protein and complex protein mixture. For peptide, A205, A280, Lowry and Ninhydrin methods had a comparable LOD (0.006 mg/ml), but Ninhydrin method had the widest detectability range (LOD-UOD=0.006-100mg/ml). For amino acid, A205 and Ninhydrin methods had a comparable LOD (0.006 mg/ml), but A205 had a wider detectability range (LOD-UOD=0.006-6.250 mg/ml). Biuret method offered the widest linear dynamic range for purified protein and complex protein mixture (0.391-100mg/ml), A280 offered the widest linear dynamic range for peptide (0.024-6.250 mg/ml), and Ninhydrin method offered the widest linear dynamic range for amino acid (0.024-0.195 mg/ml). Both Laemmli's and 2-D lysis buffers had dramatic interfering effects on all assays. Concerning the practicality and unit costs, A205 and A280 were the most favorable. Among the colorimetric methods, Bradford method consumed the least amount of samples and shortest analytical time with the lowest unit cost. These are the most extensive comparative data of commonly used quantitative protein assays that will be useful for selecting the most suitable method for each study.