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      SARS-CoV-2 RNA and antibody detection in breast milk from a prospective multicentre study in Spain

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          Abstract

          Objectives

          To develop and validate a specific protocol for SARS-CoV-2 detection in breast milk matrix and to determine the impact of maternal SARS-CoV-2 infection on the presence, concentration and persistence of specific SARS-CoV-2 antibodies.

          Design and patients

          This is a prospective, multicentre longitudinal study (April–December 2020) in 60 mothers with SARS-CoV-2 infection and/or who have recovered from COVID-19. A control group of 13 women before the pandemic were also included.

          Setting

          Seven health centres from different provinces in Spain.

          Main outcome measures

          Presence of SARS-CoV-2 RNA in breast milk, targeting the N1 region of the nucleocapsid gene and the envelope (E) gene; presence and levels of SARS-CoV-2-specific immunoglobulins (Igs)—IgA, IgG and IgM—in breast milk samples from patients with COVID-19.

          Results

          All breast milk samples showed negative results for presence of SARS-CoV-2 RNA. We observed high intraindividual and interindividual variability in the antibody response to the receptor-binding domain of the SARS-CoV-2 spike protein for each of the three isotypes IgA, IgM and IgG. Main Protease (MPro) domain antibodies were also detected in milk. 82.9% (58 of 70) of milk samples were positive for at least one of the three antibody isotypes, with 52.9% of these positive for all three Igs. Positivity rate for IgA was relatively stable over time (65.2%–87.5%), whereas it raised continuously for IgG (from 47.8% for the first 10 days to 87.5% from day 41 up to day 206 post-PCR confirmation).

          Conclusions

          Our study confirms the safety of breast feeding and highlights the relevance of virus-specific SARS-CoV-2 antibody transfer. This study provides crucial data to support official breastfeeding recommendations based on scientific evidence.

          Trial registration number NCT04768244.

          Abstract

          ARS-CoV-2 antibodies but not RNA are found in breast milk from COVID-19 infected mothers.

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          Most cited references33

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          The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments.

          Currently, a lack of consensus exists on how best to perform and interpret quantitative real-time PCR (qPCR) experiments. The problem is exacerbated by a lack of sufficient experimental detail in many publications, which impedes a reader's ability to evaluate critically the quality of the results presented or to repeat the experiments. The Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines target the reliability of results to help ensure the integrity of the scientific literature, promote consistency between laboratories, and increase experimental transparency. MIQE is a set of guidelines that describe the minimum information necessary for evaluating qPCR experiments. Included is a checklist to accompany the initial submission of a manuscript to the publisher. By providing all relevant experimental conditions and assay characteristics, reviewers can assess the validity of the protocols used. Full disclosure of all reagents, sequences, and analysis methods is necessary to enable other investigators to reproduce results. MIQE details should be published either in abbreviated form or as an online supplement. Following these guidelines will encourage better experimental practice, allowing more reliable and unequivocal interpretation of qPCR results.
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            A serological assay to detect SARS-CoV-2 seroconversion in humans

            Here, we describe a serological enzyme-linked immunosorbent assay for the screening and identification of human SARS-CoV-2 seroconverters. This assay does not require the handling of infectious virus, can be adjusted to detect different antibody types in serum and plasma and is amenable to scaling. Serological assays are of critical importance to help define previous exposure to SARS-CoV-2 in populations, identify highly reactive human donors for convalescent plasma therapy and investigate correlates of protection.
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              SARS‐CoV‐2 Seroconversion in Humans: A Detailed Protocol for a Serological Assay, Antigen Production, and Test Setup

              Abstract In late 2019, cases of atypical pneumonia were detected in China. The etiological agent was quickly identified as a betacoronavirus (named SARS‐CoV‐2), which has since caused a pandemic. Several methods allowing for the specific detection of viral nucleic acids have been established, but these only allow detection of the virus during a short period of time, generally during acute infection. Serological assays are urgently needed to conduct serosurveys, to understand the antibody responses mounted in response to the virus, and to identify individuals who are potentially immune to re‐infection. Here we describe a detailed protocol for expression of antigens derived from the spike protein of SARS‐CoV‐2 that can serve as a substrate for immunological assays, as well as a two‐stage serological enzyme‐linked immunosorbent assay (ELISA). These assays can be used for research studies and for testing in clinical laboratories. © 2020 The Authors. Current Protocols in Microbiology published by Wiley Periodicals LLC. Basic Protocol 1: Mammalian cell transfection and protein purification Basic Protocol 2: A two‐stage ELISA for high‐throughput screening of human serum samples for antibodies binding to the spike protein of SARS‐CoV‐2
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                Author and article information

                Journal
                Arch Dis Child Fetal Neonatal Ed
                Arch Dis Child Fetal Neonatal Ed
                fetalneonatal
                fnn
                Archives of Disease in Childhood. Fetal and Neonatal Edition
                BMJ Publishing Group (BMA House, Tavistock Square, London, WC1H 9JR )
                1359-2998
                1468-2052
                August 2021
                19 August 2021
                19 August 2021
                : fetalneonatal-2021-322463
                Affiliations
                [1 ]departmentDepartment of Biotechnology , Institute of Agrochemistry and Food Technology-National Research Council (IATA-CSIC) , Paterna, Valencia, Spain
                [2 ]departmentDepartment of Preservation and Food Safety Technologies , Institute of Agrochemistry and Food Technology-National Research Council (IATA-CSIC) , Paterna, Valencia, Spain
                [3 ]departmentDepartment of Gynecology and Obstetrics , Hospital Universitario Dr Peset , Valencia, Spain
                [4 ]departmentDepartment of Paediatrics , Hospital Clínico Universitario de Valencia, Nutrition Research Group of INCLIVA , Valencia, Spain
                [5 ]Health Research Institute La Fe, Neonatal Research Group and University and Polytechnic Hospital La Fe, Division of Neonatology , Valencia, Spain
                [6 ]departmentDepartment of Endocrinology , Institut de Recerca Sant Joan de Déu, Hospital Sant Joan de Déu , Barcelona, Spain
                [7 ]departmentDepartment of Infectious and Imported Diseases, Paediatric Unit , Institut de Recerca Sant Joan de Déu, Hospital Sant Joan de Déu , Barcelona, Spain
                [8 ]BCNatal - Barcelona Center for Maternal-Fetal and Neonatal Medicine, Hospital Sant Joan de Déu and Hospital Clínic, University of Barcelona, CIBERER , Barcelona, Spain
                [9 ]departmentDepartment of Biochemistry and Physiology , Universidad Barcelona Facultad Farmacia , Barcelona, Spain
                [10 ]University of Zaragoza, Hospital Clínico Universitario Lozano Blesa, Zaragoza. Instituto de Investigación Sanitaria Aragón (IIS Aragón), Red de Salud Materno Infantil y del Desarrollo (SAMID) , Zaragoza, Spain
                [11 ]departmentDepartment of Gynecology and Obstetrics , University Hospital Clinic “San Cecilio” – Health Sciences Technological Park (PTS) , Granada, Spain
                [12 ]departmentDepartment of Paediatrics , University of Granada , Granada, Spain
                Author notes
                [Correspondence to ] Dr Maria Carmen Collado, IATA, Paterna-Valencia, Spain; mcolam@ 123456iata.csic.es

                CM-C and MCC are joint senior authors.

                Author information
                http://orcid.org/0000-0001-7022-661X
                http://orcid.org/0000-0003-1347-7521
                http://orcid.org/0000-0002-6204-4864
                Article
                fetalneonatal-2021-322463
                10.1136/archdischild-2021-322463
                8384494
                34417223
                344ea41b-5342-42b9-b211-924c1925c920
                © Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.

                This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited, appropriate credit is given, any changes made indicated, and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/.

                History
                : 16 May 2021
                : 12 July 2021
                Funding
                Funded by: FundRef http://dx.doi.org/10.13039/100008666, Fundació la Marató de TV3;
                Award ID: 202106
                Categories
                Original Research
                1506
                2474
                Custom metadata
                unlocked
                free

                Neonatology
                covid-19,microbiology,neonatology,global health
                Neonatology
                covid-19, microbiology, neonatology, global health

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