The current study aimed to estimate the seroprevalence of SARS-CoV-2 IgG (S-protein) antibodies along with neutralizing assay (RBD-domain) among the whole blood donors without any prior Covid-19 history or symptoms visiting Blood Centre at a Tertiary care institution, South India amidst the ongoing pandemic. During September 2020 to March 2021, 1034 whole blood donors were enrolled into the study and were screened for anti-SARS-CoV-2 IgG antibodies using Chemiluminescence assay followed by neutralizing antibodies using surrogate neutralization ELISA. The study reported seroprevalence of 49.4%, (95% CI 46.3-52.5) among whole blood donors, with test sensitivity and specificity adjusted prevalence of 54.9% (95% CI 51.5-58.3). Seroprevalence was similar across age groups, gender, voluntary/replacement donations, area of residence, ABO and Rh groups without any statistical significance. However higher IgG antibody responses were found to be elicited in the 30-45 years age group when compared with 18-29 years age group ( value 0.046). This study also analysed the mean neutralizing capacity of SARS-CoV-2 antibodies among 97 blood donors which was 71.9 (SD: + 21.03, range 15.5 to 97.3). Donor samples with SARS-CoV-2 IgG S/Co > 9.5 had significantly higher neutralising capacity (> 68%) when compared with donor samples of S/Co < 9.5 ( value 0.000). Real-time seroprevalence studies will help to know the herd immunityamong the blood donors which will assist in knowing the Covid-19 transmission dynamics, distribution of immunity levels at a particular point in time, immunity gaps, development of novel therapeutics and prioritize the vaccination programmes to high risk individuals.