Anti-Rhesus IgG monoclonal antibody, clone TB109a [Biotin]

Chinese hamster ovary (CHO) cells are the predominant cell chassis for biopharmaceutical production. Engineering cellular pathways related to cell death, metabolism, and glycosylation in CHO cells is desired but challenging. Here, we present a novel approach that exploits CRISPR-Cas13d for gene silencing and CHO cell engineering. CRISPR-Cas13d is a burgeoning system that exploits Cas13d nuclease and guide RNA (gRNA) for RNA cleavage and gene knockdown. We first showed that CRISPR-Cas13d effectively knocked down exogenous genes in CHO cell lines (K1, DG44, and DUXB11) commonly used for recombinant protein production. We next demonstrated that CRISPR-Cas13d robustly suppressed the expression of exogenous genes and various endogenous genes involved in gene amplification, apoptosis, metabolism, and glycosylation (e.g., GS, BAK, BAX, PDK1, and FUT8) in CHO cells with efficiencies ranging from 60% to 80%, simply by transient transfection. By integrating the entire CRISPR-Cas13d system with the Sleeping Beauty system and optimal gRNA design, we further improved the knockdown efficiency and rapidly generated stable cells with approximate to 80%-90% knockdown. With this approach, we knocked down FUT8 expression for >90% and significantly attenuated the IgG fucosylation. These data altogether implicated the potentials of CRISPR-Cas13d for gene regulation, glycoengineering, and cell engineering of CHO cells.

BACKGROUND Dengue fever is endemic India and the prevalence of dengue is on the rise owing to various social and economic factors. Prevalence of asymptomatic dengue infection varies widely from less than 1 % to 80 % in India. Transfusion transmissible dengue has been reported in different parts of the word. Prevalence of subclinical dengue among blood donor poses a threat to the blood supply leading to transfusion transmissible dengue. We wanted to estimate the prevalence of IgG antibodies for Dengue in the blood donor population. METHODS Six hundred and eight whole blood donors were included in the study during the period January 2017 to October 2018. Donor registration and education was done as per the national guidelines. Donors who gave a history of previous dengue or symptoms consistent with dengue were excluded from the study. Serum samples from whole blood donors were tested for IgG antibodies using ELISA technology. RESULTS Of the total of 608 donors, 602 were male donors and 55 % of the donors were in the age group 21 - 30 years. Majority (69 %) of the donors were from urban locations. Anti IgG antibodies for dengue were present in 4.14 % of donors of which 38 % of donors were in the age group 31 - 40 years. Three hundred and eighty-four donors revealed history of fever, myalgia and headache in the past one year. No statistical significance was found between fever, myalgia and the presence of IgG anti-dengue antibodies. CONCLUSIONS The subclinical or asymptomatic prevalence of dengue infection is low when compared to other studies in other parts of the country. Enquiring into donor history for history of dengue or symptoms of dengue and deferring such donors for a recommended period will prevent transfusion transmissible dengue.