Convalescent Plasma and SARS CoV 2 COVID 19 Antibodies

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Introduction
Immunity is the body’s ability to recognize nonself-antigens
and to mount a response to a pathogen by producing anti-
bodies to prevent them from causing illness or damaging the
body.
An immune system may contain innate and adaptive compo-
nents. The innate system is essentially made up of barriers that
aim to keep viruses, bacteria, parasites, and other foreign
particles out of the body, while the adaptive system is com-
posed of more advanced lymphatic cells that are programmed
to recognise self-substances and foreign-substances (antigens
such as infectious pathogens) and to react appropriately. The
adaptive immune system, also referred to as the acquired
immunity, is a subsystem of this immune system that is
composed of specialized, systemic cells (B and T cells) and
processes that eliminate foreign antigens and any toxic
molecules they may produce.
There are two types of adaptive immunity: Active and
passive.
• Active immunity: Antibodies that develop in a person’s own
immune system after the body is exposed to an antigen
through a disease or when the person get an immunization
(i.e. a flu shot). This type of immunity lasts for a long
time.
• Passive immunity: Antibodies given to a person to prevent
disease or to treat disease after the body is exposed to an
antigen.
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Convalescent Plasma and
SARS-CoV-2 (COVID-19)
Antibodies
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The novel coronavirus SARS-CoV-2, is an enveloped
positive-sense single-stranded RNA viruses that with RNA size
ranging from 26 to 32 kb are considered to possess the largest
viral RNA genome. This large RNA covered with nucleocapsid
(N) protein is held in a phospholipid bilayer and a complex of
proteins including spike protein (S) hemagglutinin-esterase
(HE) membrane (M) and envelope (E) which provide a crown-
like shape for coronavirus. The S protein consists of two
subunits: S1 and S2. The fragment located in the middle of the
S1 subunit is the receptor-binding domain (RBD) in SARS-CoV,
which binds to the host cell receptor angiotensin-converting
enzyme 2 (ACE2).
SARS-CoV RBD contains a core and a receptor-binding motif
(RBM); the RBM mediates contacts with ACE2. The surface of
ACE2 contains two virus-binding hotspots that are essential
for SARS-CoV binding.
Even though this particular virus is just about 13–14 months
pathogen and the studies of the antibody response in infected
patients are relatively new, yet the clinical values of antibody
response are now being well understood. The antibody detec-
tion offers vital clinical information during the course of SARS-
CoV-2 infection. The findings provide strong empirical support
for the routine application of serological testing in the diagnosis
and management of COVID-19 patients.
Fig. 23.1: Structure of coronavirus SARS-CoV-2

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The present data demonstrated that typical antibody
responses to acute viral infection are induced in COVID-19
patients. To be expected, the total antibody was first detected,
followed by IgM and IgG. The seroconversion rate and the
antibody levels increased rapidly during the first two weeks,
the cumulative seropositive rate reached 50% on the 11–12 days
and 100% on the 38–40 days.43
NEUTRALIZING AND NON-NEUTRALIZING ANTIBODIES
Serological tests are valuable particularly in epidemiologic
studies and determining the immunity level of affected popula-
tion including both diagnosed and asymptomatic individuals.
Such studies also provide the primary platform for evaluation
of the hypothesis like “herd immunity”.44
Neutralizing antibodies (NAbs) are crucial in virus and
bacterial clearance and have been considered essential in
protecting against or preventing the spread of many infectious
diseases. Passive immunity driven by CBP can provide these
NAbs that restrain the infection. The RBD of the SARS-CoV-2
S protein is a major target for the antibody response. NAbs
targeting SARS-CoV RBD generally prevent the attachment of
the virus to the host cell by interfering with the binding of viral
RBD to cellular ACE2 receptor.
Non-neutralizing antibodies (non-NAbs), also referred to
as binding antibodies that bind to the pathogen but do not affect
its capacity to replicate, might contribute to prophylaxis and/
or recovery improvement. Moreover, other antibody-mediated
pathways such as complement activation, antibody-dependent
cellular cytotoxicity and/or phagocytosis may also promote
the therapeutic effect of CBP.3 In some studies it has been found
that severe COVID-19 patients had a large amount of non-
neutralizing antibodies, which may contribute to antibody-
dependent enhancement of infection.
Further Understanding of SARS-CoV-2
Neutralizing Antibodies46–49
Extensive efforts have been made on understanding antibody
recognition of the RBD at the molecular level. As of October

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2020, structures of 22 different human antibodies in complex
with the SARS-CoV-2 spike were available in the protein data
bank (PDB). Three major epitopes on the RBD are recognized
by the currently characterized SARS-CoV-2 neutralizing anti-
bodies. Most of these antibodies are specific to SARS-CoV-2, but
a few can also cross-react between SARS-CoV-2 and SARS-CoV.
These antibodies can be clustered based on their interaction
with three distinct RBD binding sites:
1. Receptor binding site;
2. CR3022 cryptic site; and
3. S309 pro-teoglycan site
Most of the antibodies that bind to these three sites show
neutralizing activity with SARS-CoV-2. Moreover, antibodies
that bind the CR3022 and S309 sites have a tendency to cross-
react with SARS-CoV RBD, as compared to antibodies to the
RBS, which are highly specific for SARS-CoV-2. Nevertheless,
recent studies have indicated that the mutation do not change
the antigenicity of the S protein and may even increase the
susceptibility of the SARS-CoV-2 to antibody neutralization.
However, some minor natural variants with an occurrence
frequency of <1%, including A475V, L452R, V483A, F490L, and
H519P on the RBD, have been shown to alter antigenicity, as
assessed by both monoclonal antibodies and convalescent sera.
Escape mutations to monoclonal antibodies have also been
identified by in vitro selection. These observations suggest that
SARS-CoV-2 could undergo antigenic drift in the future if it
becomes a seasonal virus.
Since the publication of the first structure of antibody in com-
plex with SARS-CoV-2 RBD about an year ago, many other
antibody structures that target the SARS-CoV-2 RBD have now
been determined. This compendium of structures has provided
important insights into the antigenicity and main sites of
vulnerability on SARS-CoV-2. However, it remains to be seen
whether all of the epitopes on the RBD have yet been identified
and whether regions that are not currently known to be
targeted by antibodies are truly non-immunogenic or elicit
antibodies less frequently. Since most neutralizing antibodies
to SARS-CoV-2 target the immunodominant RBD, structural
studies have been focused so far on RBD-targeting antibodies.

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It is remarkable how much structural information has already
been amassed in such a relatively short time and a testament
to prior investment in technologies, techniques and researchers
by funding agencies and our institutions over the years for
investigation of microbial pathogens.
Antibody Response to SARS-CoV-2 and Role of Neutralizing
Antibodies in COVID Convalescent Plasma
In patients with SARS-CoV infection, B cell responses typically
arise first against the nucleocapsid (N) protein. Within 4–8 days
after symptom onset, antibody responses to S protein are
found.49,50 Neutralizing antibody responses, primarily to the
S (RBD) protein, begin to develop by week 2, and most patients
develop neutralizing antibodies by week 3.51,52 Once such
patients have recovered antibodies are likely to be effective
against SARS-CoV-2. Plasma from donors who have recovered
from COVID-19 contains neutralizing antibodies to SARS-
CoV-2 virus that may help suppress the virus and modify the
inflammatory response. Convalescent plasma has been applied
with apparently good clinical results in COVID-19 and was
also previously used successfully in the treatment of SARS.53
While mechanistic correlates of protection have not yet been
identified in humans, neutralization of the virus is presumed
Fig. 23.2: Passively transferred neutralising antibodies react with spike
protein of the virus. Antibody coated viruses form clusters gets recognised
and phagocytosed

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to be an important mechanism of action for antibodies, although
the specific titre and specificity of the antibody repertoire
required (for protection) remain debatable. In SARS-CoV-2, the
primary target of neutralizing antibodies is the RBD, compris-
ing 33 amino acids in the region (amino acids 460–492) amino
acid region in the S protein, which can independently bind to
the host target ACE2 receptor.54
Acquisition and Convalescent Plasma (CP) Use
The convalescent donors must undergo standard pre-donation
assessment to ensure compliance with current regulations regard-
ing plasma donation.55 Currently, convalescent donors between
18 and 60 males and nulliparous females with weight >50 kg are
considered as subjects without infectious symptomatology for
COVID-19 after 14 days of recovery. The diagnosis of COVID- 19
infection can be either by COVID-19 RT-PCR or rapid antigen
tests. IgG antibody against COVID-19 titre of 1:640 (ELISA)
or 13 AU/mL (CLIA) or neutralizing antibodies 1:80 (PRNT/
MNT) are recommended for donor selection. As CP production
requires high quality standards, it must be free of any infection,
so tests for human immunodeficiency virus (HIV), hepatitis B,
hepatitis C, syphilis and malaria should be carried out.
Apheresis is the recommended procedure to obtain plasma.
This procedure is based on a continuous centrifugation of blood
from donor to allow selective collection plasma. The efficiency
of this technique is around 400–500 mL from a single apheresis
donation. This amount of plasma could be stored in units of
200 or 250 mL, and frozen within 24 hours of collection to be
used in further transfusions.
There is no standard transfusion dose of CP. In different
studies for coronaviruses the administration of CP ranges
between 200 and 500 mL in single or double scheme dosages.
Currently, the recommendation is to administrate 4–13 mL/kg
per dose (usually 200 mL single dose) given slowly.