However, due to the non-availability of a BSL-3 laboratory within the country, it was not possible to compare HAT with virus neutralisation in Sri Lankan individuals, and therefore the assay was compared with the sVNT, which evaluates ACE2 blocking antibodies and offers been shown to correlate well with neutralizing antibodies (Tan?et?al., 2020). the sVNT, and the level of sensitivity and persistence of antibodies in individuals with varying severity of illness was assessed inside a cohort of 71 individuals at 4C6 weeks and 13C16 weeks. The kinetics were assessed in the 1st, second, and third weeks in individuals with varying severity of acute illness. Results The specificity of the HAT was >99%, and level of sensitivity was similar to the sVNT. The levels of HAT were significantly and positively correlated with those of the sVNT (Spearman’s < 0.0001). Individuals with moderate and severe illness experienced higher HAT titres when compared to those with slight illness. Six of seven individuals with severe illness experienced a titre of >1:640 during the second week of illness, whereas only five of 31 individuals with a slight illness experienced a titre of >1:160 in the second week of illness. Conclusions Since the HAT is a simple and very cheap assay to perform, it would be ideal to use as an indication of NAbs in resource-poor settings. KEYWORDS: COVID-19, Disease severity, Haemagglutination assay, Surrogate neutralization assay, Neutralizing antibodies 1.?Intro There are several antibody assays currently in use to determine IgG, IgM, and Tetrahydrozoline Hydrochloride IgA specific to Tetrahydrozoline Hydrochloride Tetrahydrozoline Hydrochloride severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the disease responsible for coronavirus disease 2019 (COVID-19) (Algaissi?et?al., 2020; Sun?et?al., 2020; Vogelzang?et?al., 2020). While some of these assays measure total antibodies to primarily the spike protein receptor binding website (RBD), others measure IgM or IgG reactions to S1, S2, or the nucleocapsid protein (Sun?et?al., 2020; Vogelzang?et?al., 2020). These assays are sometimes used in conjunction with PCR assays, and some are used to detect those who have experienced an asymptomatic illness and in serosurveillance studies (Galipeau?et?al., 2020). Although RT-PCR is the platinum standard to identify folks who are acutely infected with SARS-CoV-2, serological checks can contribute by providing more accurate estimations of past SARS-CoV-2 infection and the immune status of the population (Ghaffari?et?al., 2020). However, while different antibody assays can be used to determine those who have been exposed to the disease, only assays that measure, or correlate with, neutralizing antibodies (NAbs), are likely to provide evidence of antibodies that are more likely to protect individuals against reinfection (Galipeau?et?al., 2020). NAbs against SARS-CoV-2 are primarily produced against the RBD of the viral spike protein (Kreer?et?al., 2020; Ni?et?al., 2020). The gold standard for determining NAbs is the plaque reduction neutralization test (PRNT), which requires biosafety level 3 (BSL-3) facilities and is time-consuming (Galipeau?et?al., 2020). With COVID-19 distributing at an exponential rate around the world, many assays have been developed to measure NAbs that can be performed within a few hours inside a BSL-2 facility (Tan?et?al., 2020; Townsend?et?al., 2020). One such assay is the surrogate disease neutralization test (sVNT), which actions the percentage of inhibition of binding of the RBD of the spike protein to recombinant angiotensin-converting enzyme 2 (ACE2) (Tan?et?al., 2020). Further, Townsend et?al. have developed a haemagglutination test (HAT) to measure antibodies to the RBD, where the RBD of the disease is linked to a nanobody IH4, specific for any conserved epitope within glycophorin A on reddish blood cells (RBCs) (Townsend?et?al., 2020). In the presence of antibodies to RBD, IH4-RBD-6H bound RBCs will display agglutination. Since the majority of NAbs SLC2A2 are directed at the RBD (Kreer?et?al., 2020; Ni?et?al., 2020) and the level of antibodies detected from the HAT correlates with the neutralising half maximal?inhibitory concentration (IC50)(manuscript in preparation), this assay could be used as an inexpensive test to predict NAbs in study and community settings where high throughput assays are required (Townsend?et?al., 2020). Consequently, in order to determine the usefulness of the HAT, the performance of the HAT was compared with that of an existing assay that is used like a surrogate to measure the presence of NAbs by blockade of ACE2 binding from the RBD. 2.?Methods 2.1. The HAT to detect neutralizing antibodies The HAT was performed as explained previously (Townsend?et?al., 2020). Briefly, RBCs from an O bad donor were mixed with the IH4-RBD-6H (a nanobody against a conserved glycophorin A epitope on RBCs, linked to the RBD of SARS-CoV-2) and incubated for 1?hour with serum. Phosphate buffered saline (PBS) was used as a.