Biology subjects

Hopp, C. S.

Publications and source records attributed to Hopp, C. S..

4 recordsLinked to original sources

bioRxiv2022

Application of B cell immortalization for the isolation of antibodies and B cell clones from vaccine and infection settings

The isolation and characterization of neutralizing antibodies from infection and vaccine settings will inform future vaccine design, and methodologies that streamline the isolation of antibodies and the generation of B cell clones are of great interest. Retroviral transduction to express Bcl-6 and Bcl-xL in primary B cells has been shown to promote long-term B cell survival and antibody secretion in vitro, and can be used to isolate antibodies from memory B cells. The application of this methodology to B cell subsets from tissues and to B cells from individuals with chronic infection has not been extensively characterized. Here, we characterize Bcl-6/Bcl-xL B cell immortalization across multiple tissue types and B cell subsets in healthy and HIV-1 infected individuals, as well as individuals recovering from malaria. In HIV-1- and malaria-uninfected donors, naive and memory B cell subsets from PBMC and tonsil tissue transformed with similar efficiencies, and displayed similar characteristics after transformation with respect to their longevity and immunoglobulin secretion. In HIV-1-viremic individuals or in individuals after malaria infection, the CD27-CD21- memory B cell subsets transformed with lower efficiencies compared to the CD27+CD21+ populations, but following transformation B cells expanded and secreted IgG with similar efficiency. Using B cells from HIV-1-infected individuals, we combined Bcl-6/Bcl-xL B cell immortalization with a HIV-1 microneutralization assay to isolate broadly neutralizing antibodies related to VRC13 and VRC38.01. Overall, Bcl-6/Bcl-xL B cell immortalization can be used to isolate antibodies and generate B cell clones from multiple different B cell populations, albeit with different efficiencies.

immunology↗

bioRxiv2021

Atypical B cells upregulate co-stimulatory molecules during malaria and secrete antibodies with T follicular helper cell support

Several infectious and autoimmune diseases are associated with an expansion of CD21-CD27- atypical B cells (atBCs). The function of atBCs remains unclear and few studies have investigated the biology of pathogen-specific atBCs during acute infection. Here, we performed longitudinal RNA-sequencing and flow cytometry analyses of Plasmodium falciparum (Pf)-specific B cells before and shortly after febrile malaria, with simultaneous analysis of influenza hemagglutinin (HA)-specific B cells as a comparator. B cell receptor-sequencing showed that Pf-specific atBCs, activated B cells (actBCs) and classical memory B cells share clonality and have comparable somatic hypermutation. In response to malaria, Pf-specific atBCs and actBCs expanded and upregulated molecules that mediate B-T cell interactions, suggesting that atBCs respond to T follicular helper (Tfh) cells. Indeed, in the presence of Tfh cells and Staphylococcal enterotoxin B, atBCs of malaria-exposed individuals differentiated into CD38+ antibody-secreting cells in vitro, suggesting that atBCs may actively contribute to humoral immunity to infectious pathogens. One Sentence SummaryThis study shows that atypical B cells actively respond to acute malaria and have the capacity to produce antibodies with T cell help.

immunology↗

bioRxiv2019

Quantitative intravital imaging of Plasmodium falciparum sporozoites: A novel platform to test malaria intervention strategies

Malaria infection starts with the injection of motile Plasmodium sporozoites into the hosts skin during a mosquito bite. Previous studies using the rodent malaria model indicate that the dermal inoculation site may be where sporozoites are most vulnerable to antibodies, yet, functional in vivo assays with human malaria parasites are lacking. Here, we present the first characterization of P. falciparum sporozoites in the skin, comparing their motility to two rodent malaria species and investigating whether the environment of its natural host influences P. falciparum sporozoite motility using a human skin xenograft model. The combined data suggest that in contrast to the liver and blood stages, the skin is not a species-specific barrier for Plasmodium. We observe that P. falciparum sporozoites inoculated into mouse skin move with similar speed, displacement and duration, and enter blood vessels in similar numbers as the rodent parasites. Thus, interventions targeting P. falciparum sporozoite migration can be tested in the murine dermis. Importantly, to streamline quantification of sporozoite motility, we developed a toolbox allowing for automated detection and tracking of sporozoites in intravital microscopy videos. This establishes a platform to test vaccine candidates, immunization protocols, monoclonal antibodies and drug candidates for their impact on human malaria sporozoites in vivo. Screening of intervention strategies for in vivo efficacy against Pf sporozoites using this new platform will have the potential to validate targets prior to expensive clinical trials.

microbiology↗

bioRxiv2018

Generation of Transgenic Human Malaria Parasites With Strong Fluorescence in the Transmission Stages

Malaria parasites have a complex life cycle that includes specialized stages for transmission between their mosquito and human hosts. These stages are an understudied part of the lifecycle yet targeting them is an essential component of the effort to shrink the malaria map. The human parasite Plasmodium falciparum is responsible for the majority of deaths due to malaria. Our goal was to generate transgenic P. falciparum lines that could complete the lifecycle and produce fluorescent transmission stages for more in-depth and high-throughput studies. Using zinc-finger nuclease technology to engineer a marker-free integration site, we generated three transgenic P. falciparum lines in which tdtomato or gfp were stably integrated into the genome. Expression was driven by either stage-specific peg4 and csp promoters or the constitutive ef1a promoter. Phenotypic characterization of these lines demonstrates that they complete the life cycle with high infection rates and give rise to fluorescent mosquito stages. The transmission stages are sufficiently bright for intra-vital imaging, flow cytometry and scalable screening of chemical inhibitors and potentially inhibitory antibodies.

microbiology↗