Acanthamoeba spp. monoclonal antibody against a CPA2 transporter: a promising molecular tool for acanthamoebiasis diagnosis and encystment study

Abstract

Free-living amoeba of the genus Acanthamoeba are ubiquitous protozoa involved in opportunistic and non-opportunistic infection in humans, such as granulomatous amoebic encephalitis and amoebic keratitis. Both infections have challenging characteristics such as the formation of the resistant cysts in infected tissues, hampering the treatment and most usual diagnosis depending on time-consuming and/or low sensitivity techniques. The use of monoclonal antibodies presents itself as an opportunity for the development of more effective alternative diagnostic methods, as well as an important and useful tool in the search for new therapeutic targets. This study investigated the possibility of using a previously produced monoclonal antibody (mAb3), as a diagnostic tool for the detection of Acanthamoeba trophozoites by direct and indirect flow cytometry and immunofluorescence. Immunoprecipitation assay and mass spectrometry allowed the isolation of the antibody's target and suggested it is a transporter part of the CPA (cation: proton antiporter) superfamily. In vitro tests indicate an important role of this target in Acanthamoeba's encystment physiology. Our results support the importance of studying the role of CPA2 transporters in the context of acanthamoebiasis, as this may be a way to identify new therapeutic candidates.

Characterization of a Peptide Antibody Specific to the Adenylyl Cyclase-Associated Protein of Acanthamoeba castellanii

“…Accordingly, sensitive and rapid differential diagnostic methods for AK have been stud-ied. Currently, several studies investigating the diagnostic potential of antibody-based methods for AK have been reported [7][8][9][10][11]. In this study, we produced a polyclonal peptide antibody against the ACAP of A. castellanii and studied its specificity.…”

“…Immunoassays using specific antibodies are also important techniques for the detection of Acanthamoeba. Four monoclonal antibodies (AMEC1, AMEC3, MTAC1, and MTAC3) generated by mixtures of A. castellanii trophozoites and cysts, a monoclonal antibody against a CPA2 transporter of A. castellanii, and a monoclonal antibody against a mannose-binding protein of A. culbertsoni have been produced to support AK diagnosis [7][8][9]. Polyclonal antibodies against inosine-uridine preferring nucleoside hydrolase (IPNH) and chorismate mutase (CM) of A. castellanii have also been produced for specific detection of Acanthamoeba [10,11].…”

Monoclonal Antibodies and Invasive Aspergillosis: Diagnostic and Therapeutic Perspectives

“…Monoclonal antibodies (mAbs), produced by a single B-lymphocyte clone, have high specificity in targeting the corresponding antigen(s) and have the potential to improve diagnostic tests, forming the basis for novel IA treatments. MAbs have been developed for diagnosis and therapy of cancers [ 11 , 12 ], autoimmune disease [ 13 ], asthma [ 14 ] and infectious diseases [ 15 , 16 ] including COVID-19 [ 17 , 18 ] but have yet to reach their full potential for improving the outcomes of IA. This review examines the potential usefulness of mAbs in diagnosing and treating IA from different perspectives.…”

A history of over 40 years of potentially pathogenic free-living amoeba studies in Brazil - a systematic review

“… 41 , 42 , 43 Additional targets for detection purposes are cation:proton antiporter protein family (CPA) and a calreticulin extracellular domain. 44 , 45 The former was identified through the monoclonal antibody mAb3, which positively reacted with several Acanthamoeba pathogenic strains. 44 The latter study described the use of a recombinant polypeptide, produced from a predicted sequence of A. castellanii calreticulin, which was recognised by sera from rats with GAE in the ELISA test.…”