Biology subjects

Marchi, S.

Publications and source records attributed to Marchi, S..

3 recordsLinked to original sources

bioRxiv2023

Establishment and validation of a High-throughput Micro-Neutralization assay for Respiratory Syncytial Virus (subtypes A and B)

The validation of a bioanalytical method allows us to determine its validity for a designated purpose and to guarantee the reliability of its analytical results. The virus neutralization assay has proved to be suitable for the detection and quantization of specific serum neutralizing antibodies against respiratory syncytial virus subtypes A and B. Respiratory syncytial virus is a negative-sense RNA virus and is responsible for the majority of acute lower respiratory tract infections in infants and older adults worldwide. Owing to its widespread infection, the WHO considers it a target for the development of preventive vaccines. Despite the high impact of its infections, however, no vaccine candidate is currently available. The aim of this paper is to provide a detailed validation process for the micro-neutralization assay and to demonstrate that this method can effectively support the efficacy assessment of candidate vaccines and the definition of correlates of protection.

immunology↗

bioRxiv2020

Comparative analyses of SARS-CoV-2 binding (IgG, IgM, IgA) and neutralizing antibodies from human serum samples

A newly identified coronavirus, named SARS-CoV-2, emerged in December 2019 in Hubei Province, China, and quickly spread throughout the world; so far, it has caused more than 18 million cases of disease and 700,000 deaths. The diagnosis of SARS-CoV-2 infection is currently based on the detection of viral RNA in nasopharyngeal swabs by means of molecular-based assays, such as real-time RT-PCR. Furthermore, serological assays aimed at detecting different classes of antibodies constitute the best surveillance strategy for gathering information on the humoral immune response to infection and the spread of the virus through the population, in order to evaluate the immunogenicity of novel future vaccines and medicines for the treatment and prevention of COVID-19 disease. The aim of this study was to determine SARS-CoV-2-specific antibodies in human serum samples by means of different commercial and in-house ELISA kits, in order to evaluate and compare their results first with one another and then with those yielded by functional assays using wild-type virus. It is important to know the level of SARS-CoV-2-specific IgM, IgG and IgA antibodies in order to predict population immunity and possible cross-reactivity with other coronaviruses and to identify potentially infectious subjects. In addition, in a small sub-group of samples, we performed a subtyping Immunoglobulin G ELISA. Our data showed an excellent statistical correlation between the neutralization titer and the IgG, IgM and IgA ELISA response against the receptor-binding domain of the spike protein, confirming that antibodies against this portion of the virus spike protein are highly neutralizing and that the ELISA Receptor-Binding Domain-based assay can be used as a valid surrogate for the neutralization assay in laboratories which do not have Biosecurity level-3 facilities.

immunology↗

bioRxiv2018

NHBA is processed by kallikrein from human saliva

Neisserial Heparin Binding Antigen (NHBA) is a surface-exposed lipoprotein and a component of the Bexsero vaccine. NHBA is characterized by the presence of a highly conserved Arg-rich region involved in binding to heparin and heparin sulphate proteoglycans present on the surface of host epithelial cells, suggesting a possible role of NHBA during N. meningitidis colonization. NHBA has been shown to be cleaved by the bacterial NalP protein, a meningococcal protease and by human lactoferrin (hLF), a host protease present in different body fluids (saliva, breast milk and serum). Cleavage occurs upstream or downstream the Arg-rich region. Since the human nasopharynx is the only known reservoir of infection, we further investigated the susceptibility of NHBA to human proteases present in the saliva to assess whether proteolytic cleavage could happen during the initial steps of colonization. Here we show that human saliva proteolytically cleaves NHBA; and identified human kallikrein 1 (KLK1) as the main protease responsible for this cleavage. Kallikrein is an important enzyme present in blood plasma, lymph, urine, saliva, pancreatic juices, and other body fluids that catalyze the proteolysis of several human proteins. We report the in vitro characterization of NHBA cleavage by kallikrein; the identification of the cleavage in the recombinant NHBA protein and, on the native protein, when expressed on live bacteria. Overall, this findings provide new insights on NHBA as target of host proteases, highlights a potential role of NHBA in the Neisseria meningitidis nasopharyngeal colonization, and of kallikrein as a defensive agent against meningococcal infection.

biochemistry↗