Generation and Characterization of Anti-chitinase Monoclonal Antibodies

Abstract

Class IV chitinase, an allergenic protein of Vitis vinifera (grape), was purified by anion exchange chromatography and used for immunization of Balb/c mice. Monoclonal antibodies (MAbs) were raised by hybridoma technology using Sp2/0 myeloma cells. Finally after three limiting dilutions, six stable clones were generated. Antibody isotyping showed that IgG(2a), IgG(2b), and IgM were produced by one, two, and three of the clones, respectively. All of the MAbs had kappa light chain. The affinities were in the range of 3 × 10(8) to 1.2 × 10(9) M(-1). The MAbs were specific for grape chitinase as confirmed by Western blotting. In conclusion, we successfully produced several MAbs against grape class IV chitinase, which could be used for assessment of this allergen in different grape cultivars.

Graphene Oxide Attenuates Th2-Type Immune Responses, but Augments Airway Remodeling and Hyperresponsiveness in a Murine Model of Asthma

“…This clearly indicates that the presence of GO or factors such as chitinases that are released upon GO stimulation during an allergen sensitization triggers such responses. This notion is further supported by previous immunization studies in mice. , Immunizations using chitinases in the presence of adjuvants such as aluminum, as used in this study, elicited strong IgG responses including IgG2a. , The use of chitosan-based adjuvants during vaccination also induced significantly enhanced levels of IgG1 and IgG2a in the serum . Further, mice immunized with Derf2-47-67-loaded chitosan particles had increased levels of Der f-specific IgG2a in the serum .…”

“…This notion is further supported by previous immunization studies in mice. 59,60 Immunizations using chitinases in the presence of adjuvants such as aluminum, as used in this study, elicited strong IgG responses including IgG2a. 59,60 The use of chitosan-based adjuvants during vaccination also induced significantly enhanced levels of IgG1 and IgG2a in the serum.…”

“…59,60 Immunizations using chitinases in the presence of adjuvants such as aluminum, as used in this study, elicited strong IgG responses including IgG2a. 59,60 The use of chitosan-based adjuvants during vaccination also induced significantly enhanced levels of IgG1 and IgG2a in the serum. 46 Further, mice immunized with Derf2-47-67-loaded chitosan particles had increased levels of Der f-specific IgG2a in the serum.…”

Expression of recombinant parvalbumin from wolf-herring fish and determination of its IgE-binding capability

“…We also included sera from seven non-allergic healthy individuals who declared no clinical symptoms following fish consumption and whose sera showed no significant immunoreactivities with fish mix ELISA disks as negative controls (detailed demographic data published in Mohammadi et al, 2016). We also immunized two female BALB/c mice with native protein as described elsewhere (Soukhtanloo, Falak, Sankian, & Varasteh, 2011), and their sera were collected and used to evaluate IgG antibodies. The study was approved by the Medical Ethics Committee of Tehran University of Medical Sciences, Tehran, Iran.…”

“…For evaluation of the IgG reactivity of the recombinant parvalbumin, we used sera from native parvalbumin-sensitized mice. For this purpose, a couple of female BALB/c mice were immunized with subcutaneous administration of 100 μg of native parvalbumin and twice subsequent 50 μg boosters with 2-week intervals as previously described (Soukhtanloo et al, 2011). Then sera were collected and used for evaluation of parvalbumin-specific IgG reactivity and inhibition studies.…”