**1. Introduction**

According to the WHO, the COVID-19 pandemic, caused by a new pathogen SARS-CoV-2 in the human population, which began at the end of 2019 in Wuhan, in China's Hubei province, has affected 220 countries and territories to date. As of 29 June 2021, the total number of reported cases worldwide exceeds 180 million, and the number of deaths worldwide is almost 4 million [1]. In many countries, quarantines and other milder strategies for preventing the spread of the infection, such as physical distancing and the use of masks, may have prevented most people from being infected. According to the opinion of some researchers, this is a paradox, as such measures leave people without immunity, susceptible to new waves of infection. Healthcare workers, the elderly, and people with medical conditions, such as cardiovascular and cerebrovascular diseases, diabetes, and neoplasms, are at a particularly high risk of infection [2,3]. It is quite possible that the

**Citation:** Kazachinskaia, E.; Chepurnov, A.; Shcherbakov, D.; Kononova, Y.; Saroyan, T.; Gulyaeva, M.; Shanshin, D.; Romanova, V.; Khripko, O.; Voevoda, M.; et al. IgG Study of Blood Sera of Patients with COVID-19. *Pathogens* **2021**, *10*, 1421. https://doi.org/10.3390/ pathogens10111421

Academic Editors: Philipp A. Ilinykh, Kai Huang and Xuguang Li

Received: 15 July 2021 Accepted: 30 October 2021 Published: 2 November 2021

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**Copyright:** © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

modern world will not return to "pre-pandemic normality" until safe and effective vaccines have been developed and a global vaccination program has been implemented [4].

The latest reports show that most COVID-19 patients develop lymphopenia as well as pneumonia, with higher plasma levels of pro-inflammatory cytokines in severe cases, suggesting that the host immune system is involved in the pathogenesis [5]. However, there is still a very limited understanding of the immune responses, especially adaptive immune responses to SARS-CoV-2 infection. Since COVID-19 is a new disease for humanity, and the nature of the protective immune responses against it is poorly understood, it is also unclear what vaccination strategies will be the safest and most immunologically successful. It is quite possible that vaccine-induced protection may differ from natural immunity due to the virus's different evading strategies [6] and/or due to acquired humoral immunopathology in the form of antibody-dependent enhancement of infection and/or antigenic imprinting [7]. It is important to understand adaptive immunity to SARS-CoV-2, not only for vaccine development, but for interpreting COVID-19 pathogenesis and the calibration of pandemic control measures.

In this regard, data from the studies of the natural immune response to COVID-19 will contribute to COVID-19's pathogenesis, the development of effective vaccines, and therapeutic strategies. It is especially important to make clear the difference in immune responses between asymptomatic, mild, and severe cases and in the early and late stages of infection. In addition, the fact that asymptomatic and mildly suffering people develop a low level of antibody-mediated protection is important for assessing herd immunity [6]. At this moment, in the territory of Russia, this topic is not highlighted fully enough. Our investigation relates to the study of the immune response in patients with a different course of COVID-19 at the very beginning of the pandemic in one of the biggest cities of Russia under restrictions (the absence of air and railway connections with neighboring countries). It may reflect the situation at a specific point and is valuable precisely in a retrospective analysis of the development of the epidemic process in comparison with both other regions of Russia and other countries. In detail, the aim of this study was to examine humoral immunity in the samples of convalescents' blood sera with COVID-19 in a range of severities, and patients who subsequently died in hospital from this disease, using native SARS-CoV-2 and its individual recombinant proteins to visualize an individual immune "picture" of antibodies, i.e., their profile for specificity to the N proteins, the S trimer, and RBD. We consider the presence of such antibodies as part of the immune response to SARS-CoV-2. Previously, it was reported that sera from some patients could inhibit SARS-CoV-2 entry in target cells, indicating the involvement of humoral immunity due to anti-S antibodies as early as three days post symptom onset. Protein N is a major immunogen, and antibodies to it are formed in some patients who have been ill [5]. The antibodies to protein M are observed less often, and we assume that this fact would also be interesting to study. The cross-activity of antibodies with the inactivated SARS-CoV antigen (2002) was also assessed.

## **2. Results**

To study humoral immunity against SARS-CoV-2, a random sample of 54 blood sera received from 26 convalescents (1 asymptomatic case, 13 mild cases, 1 moderate case with hospitalization for pneumonia and 1 without hospitalization, 10 severe cases, and 13 severe cases with a lethal outcome in the hospital) was used. In some cases, serum was paired from one patient. The titers of specific interaction of antibodies with these antigens were found in blood sera by the author's laboratory ELISA, using inactivated wholecell SARS-CoV-2/human/RUS/Nsk-FRCFTM-1/2020 and SARS-CoV (strain Frankfurt 1, 2002) preparations as antigens, as well as the SARS-CoV-2 recombinant proteins—the N nucleoprotein, the full S trimer (spike) and RBD, its receptor-binding domain in the S1 region. For recombinant proteins, preparation the plasmids for the expression of the full N and full S trimer (spike) and also its part in the S1 region, RBD, were constructed with further purification of obtained proteins.
