Neutralizing activity to SARS-CoV-2 of convalescent and control plasma used in a randomized controlled trial

Abstract

Background: There are limited data on the neutralizing activity of convalescent plasma (CP) administered in randomized controlled trials (RCT) of COVID-19 infection.

Study design and methods: As part of an RCT, CP was collected per FDA guidelines from individuals recovered from COVID-19 infection. CP donors had to have ≥145 optical density (OD) units (ideal target ≥300) using a semiquantitative, immunochromatographic test for IgG antibody to the nucleocapsid protein (NP) of SARS-CoV-2 (typical range 0-500 OD units). A random subset of samples [14 control plasma, 12 CP "medium-anti-NP" (145-299 OD units), and 13 CP "high" anti-NP (≥300 OD units)] were tested for neutralizing antibodies using an established viral luciferase antibody inhibition assay to detect the infection of SARS-CoV-2 pseudovirus that encoded spike protein (SARS2-S_trunc ) on a human immunodeficiency virus 1 vector (NL43dEnvNanoLuc), using ACE2-expressing 293 T cells. The titer needed to neutralize 50% of viral activity (NT50) was calculated.

Results: The uptake of SARS-CoV-2 pseudovirus by 293T_ACE2 cells was inhibited by pretreatment with CP compared to control CP (p < .001) with control plasma having a median (IQR) 50% neutralization titer (NT50) of 1:28 (1:16,1:36) compared to 1:334 (1:130,1:1295) and 1:324 (1:244,1:578), for medium anti-NP and high anti-NP CP units, respectively. The neutralizing activity of CP met minimum FDA criteria with neutralizing antibody titers >1:80 in 100% of randomly selected samples, using a conservative approach that excluded non-specific binding.

Discussion: Plasma from donors screened using an immunochromatographic test for IgG antibody to SARS-CoV-2 NP exhibited neutralizing activity meeting FDA's minimum standard in all randomly selected COVID-19 CP units.

Keywords: FFP transfusion; immunology (other than RBC serology).

FIGURE 1

Inhibition of SARS‐CoV‐2 spike protein by control and convalescent plasma. Infectivity curves for SARS‐CoV‐2 pseudovirus incubated with control or convalescent plasma serially diluted between 1:4 and 1:2916. “Control” on the x‐axis represents the average luciferase activity with control plasmas at 1:4 dilution. (A) Depicts results from control plasma, 1(B) from the group of plasma possessing between 149 and 299 OD anti‐NP IgG (medium anti‐NP), and (C) from the group harboring >300 OD anti‐NP IgG (high anti‐NP). In (A)‐(C), plasma samples where an NT50 could be determined are marked with a filled in circle versus an open circle for samples for which an NT50 value could not be calculated. Aggregate curves (mean, SD) for these 3 groups are shown in (D)

FIGURE 2

Neutralizing capacity (NT50) for control and convalescent plasma. NT50 for control and convalescent plasma (medium, high anti‐NP) of CP. Each dot represents the calculated NT50 value for an individual plasma sample. The dashed lines at 1:80 correspond with the minimum titer recommended by the FDA for CP, and with the “optimally greater than 1:320” titer stated by FDA in the initial (March 24, 2020) guidance document [Color figure can be viewed at wileyonlinelibrary.com]