A total of 273 paired DBS-serum samples were analyzed, of which 129 were positive, as assessed from the research test. screening using Youden’s J index. A total of 273 combined DBS-serum samples were analyzed, of which 129 were positive, as assessed by the research test. The sensitivities and specificities of DBS screening ranged from 95.0% to 97.1% and from 97.1% to 98.8%, respectively, depending on the human population (residents or staff members) and the DBS card type. Consequently, we found that DBS sampling is definitely a valid alternative to venipuncture for the detection of SARS-CoV-2 antibodies among seniors subjects. IMPORTANCE Since the implementation of newly developed SARS-CoV-2 vaccines in the Pramiracetam general human population, serological checks are of increasing importance. Because DBS samples can be obtained having a finger prick and may become shipped and stored at space temp, they may be ideal for use in large-scale SARS-CoV-2 serosurveillance or postauthorization vaccination studies, actually in an seniors study human population. for 8 min, and the tubes were stored at 4C. The following day time, serum was aliquoted into fresh serum vials and analyzed by means of the SARS-CoV-2 IgG Architect immunoassay (Abbott Laboratories). DBS samples selected for analysis (observe below) were analyzed a maximum of 2 days after collection. SARS-CoV-2 IgG detection in serum samples by means of CMIA. All serum samples were analyzed for antinucleocapsid SARS-CoV-2 IgG serological results by using the Architect i2000sr Plus system (Abbott). This system allows high-throughput screening of the sera, providing real-time info on the number of positive samples. This way, we could rapidly validate DBS checks for our ongoing and future studies in NHs. The Architect i2000sr Plus system uses the CMIA technique to detect antibodies. After thawing the sera and vortex-mixing them briefly, the Architect system analyzes the samples instantly using a SARS-CoV-2 assay, a specific calibrator Pramiracetam kit, and a specific control kit. We used an in-house-validated cutoff index of 0.9 to classify sera as positive (0.9) or negative ( 0.9) for SARS-CoV-2 IgG antibodies. SARS-CoV-2 IgG detection in DBS samples by means of ELISA. The DBS samples were analyzed for the presence of antispike (S1 antigen) SARS-CoV-2 IgG antibodies by means of ELISA (EUROIMMUN; PerkinElmer Health Sciences Inc.) and not the Abbott CMIA because the second option assay requires a volume of 150?l, which is more than the volume of capillary blood absorbed on one circle of the DBS cards. One circle (6-mm diameter) was cut out from each DBS cards using a puncher and was placed in a well of a sterile 96-well U-shaped microtiter plate. To avoid cross-contamination, the puncher was Pramiracetam cleaned between punches using a 70% alcohol remedy and a cotton swab. A total volume of 250?l preheated (1?h at 37C) sample buffer was added to each sample well of the 96-well microtiter plate, and the plate was incubated at 37C for 1 h. After mild mixing of the eluate by means of up-and-down pipetting, a total of 100?l of this eluate was utilized for ELISA, according to the manufacturers instructions. The ELISA was run on the automated Behring Elisa Processor III (Siemens AG, Munich, Germany). DBS samples were classified according to their Rabbit Polyclonal to MAST3 antibody index OD value (450 nm) as bad ( 1.1) or positive (1.1), while recommended by the manufacturer. The borderline category was not considered. Sample size and analysis. The sample size was determined using the strategy explained by Buderer, focusing on the level of sensitivity (15). Here, we hypothesized the level of sensitivity of DBS screening would be lower than that of serum screening due to antibodies being affected by or remaining captured in the protein saver cards. Using an anticipated level of sensitivity of 85%, an level of 0.05, and a precision parameter () of 0.10, we needed a minimum of 49 positive serum samples, which were collected for both the residents and the staff members. The level of sensitivity and specificity of the DBS checks were determined using the Abbott CMIA with sera from venous blood samples as a research test. The 95% CIs were determined using the Wilson-Brown method (16). To determine the ideal cutoff value for the Whatman and EUROIMMUN DBS samples, we determined Youdens J index. The accuracy was determined using the AUC of the ROC curves. All analyses were performed using the statistical software GraphPad Prism 6 (GraphPad Software Inc., San Diego, CA, USA). ACKNOWLEDGMENTS We say thanks to all occupants and their families, staff members, and users of management in the NHs that.