Micro Analytical Immunochemistry

National Institute Of Biomedical Imaging And Bioengineering, Bethesda

Investigators

Linked publications, trials & patents

Abstract

Sub-project #1 MALDI imaging of tissue samples to identify novel proteins MALDI-TOF Imaging is an emerging tool for the label free measurement of peptides, proteins, lipids, drugs and metabolites. The tissue can be imaged directly be the MALDI-TOF and combined with other modalities to study the molecular profiles as well as spatial placement of the tissue samples. Currently most tissue samples are imaged by MALDI and compared to histological stained tissues. In collaboration with NIAID, we are developing protocols to image tissues and then compare them to confocal microscopy/histo-cytometry imaged sections. The confocal microscopy imaging provides cellular level of resolution and spatial mapping whereas MALDI-IMS provides more high-throughput proteomics and molecular mapping that can identify biomolecules associated with tissue inflammation and other processes. The goal is to use these techniques to image lymph nodes infected with HIV in hopes of identifying new biomarkers and providing spatial resolution of them. In collaboration with LDRR, we are developing protocols to image mice tissue that has been irradiated with ultrasound. We are imaging tissues pre and post treatment to determine if any proteins, peptides or lipids are unregulated by this procedure and identify them if they are. The goal of is to determine if the mice exhibit an inflammatory response to the procedure. Sub-Project #2 Trace metal analysis by Inductively Coupled Plasma (ICP) ICP analysis with UV/Vis detection can be used to analyze samples for the presence of trace metals. The samples are introduced into a plasma flame and then detected by a continuous wavelength spectrophotometer to allow for quantitative measurements of the metal of interest. This technique allows researchers to determine yields for reactions, measure cellular uptake of metal compounds or determine metal levels in tissues in contrast agent experiments. Studies using ICP-OES include: - Analysis of magneto-plasmonic Janus vesicles integrated with Gold nanoparticles - Analysis of tissue samples for the presence of Boron nanoparticles - Analysis of multimodal contrast agents to quantify the amount of iron and gadolinium present - Analysis of novel drug eluting beads to quantify the amount of boron present - Analysis of poly(acrylic acid) gels to quantify the amount of sodium, calcium, potassium and chlorine present. - Analysis of saliva collected from patients diagnosed with Hepatitis C to quantify sodium and potassium content - Analysis of saliva collected from mice modeling dry mouth syndrome Sub-project #3 High-throughput analysis of IgG glycosylation by matrix assisted laser desorption/ionization (MALDI) -time-of-flight (TOF)- mass spectrometry (MS). The Laboratory of Immunoregulation at NIAID focuses its research efforts on the elucidation of cellular and molecular mechanisms of the regulation of the human immune response in health and disease. A major component of these research efforts involves understanding the immunopathogenesis of B cells in HIV infection. Dr. Susan Moir of the B-Cell Immunology Unit has recently described a role for IgG3 in regulating B cells in HIV-infected individuals enrolled in clinical research protocol 02-I-0202. The findings indicated that glycosylation of IgG3 and possibly other serum proteins played a role in the regulatory function described in the study; the findings were obtained from the analysis of specimens isolated from over 100 participants. The evaluation of glycosylation patterns in serum proteins must be performed using an approach that is compatible with processing a relatively large number of samples from a relatively small quantity of material. Preliminary assays will be performed on a few positive and negative controls, followed by a few longitudinal samples from individuals who either begin with a positive IgG3 profile and then lose it over time or acquire the profile over time. Sub-project #4 Serosurvey to determine antibody presence to SARS-COV-2 protein The Section for Immuno-Engineering in NIBIB has undertaken the development of and ELISA based serosurvey to detect the presence of IgG, IgM and IgA antibodies to SARS-COVID2 SPIKE and RBD proteins. Studies using this serosurvey include: - 9,300 healthy blood samples were collected at time point zero to determine the extent of community spread of SARS-COVID-2 within the community. - Approximately 7000 healthy blood samples were collected from the initial individuals at time point six months to determine the extent of community spread and vaccination status. - Approximately 5000 healthy blood samples were collected from the initial individuals at time point one year to determine the extent of community spread and vaccination status. - 2,542 samples from trauma patients to determine seropositivity for SARS-COV-2 - Autopsy samples from patients suspected of SARS-COV-2 death to determine seropositivity for SARS-COV-2 - Approximately 15 samples from patients who experienced rebound infections after being administered paxlovid to determine seropositivity - Approximately 75 health care worker samples who have received boosters to determine seropositivity - Approximately 30 samples of patients enrolled in an IL-7 trial to determine seropositivity - Approximately 4000 samples of individuals with select immunodeficiencies and immune dysregulations compared to healthy volunteers to assess pre- and post- vaccine immune responses - Approximately 300 samples of individuals with Mitochondrial Disease to determine seropositivity