Micro Analytical Immunochemistry

National Institute Of Biomedical Imaging And Bioengineering, Bethesda

Investigators

Linked publications, trials & patents

Abstract

Sub-project #1 Matrix Assisted Laser Desorption Ionization (MALDI)- Time of Flight (TOF) 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 by the MALDI-TOF and combined with other modalities to study the molecular profiles as well as spatial placement of the molecules within the tissue samples. Currently most tissue samples are imaged by MALDI and compared to histological stained tissues. In collaboration with the Laboratory of Diagnostic Radiology Research, 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 upregulated by this procedure and identify them if they are. The goal of this project is to determine if the mice exhibit an inflammatory response to the procedure. In collaboration with NEI, we are developing protocols to image mouse eyes that have been dosed with small molecule therapy agents that address inherited retinal disease. The protocols will help identify the spatial distribution of these agents within the eye. Additionally, the protocols will help to identify novel lipids and peptides that are observed in eyes with this disease. In collaboration with NCI, we are developing protocols to image bone for the presence of citrate and other lipid molecules that may be indicative of prostate cancer metastasis. This protocol will expand the biological samples we analyze to include bone samples. Sub-Project #2 Trace metal analysis by Inductively Coupled Plasma (ICP)- Optical Emission Spectroscopy (OES) spectroscopy ICP analysis with UltraViolet/Visible 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 manganese oxide nanoparticles for the amount of manganese in the particle and released in acidic solutions - Analysis of saliva samples for Calcium, Sodium and Potassium from patients being treated with AAV2hAQP1 therapy - Analysis of serum, tumor and stool samples to quantify the amount of Manganese, Cobalt, Copper, Iron, Nickel and Zinc present in pediatric patients with PPGL tumors. - Analysis of saliva collected from patients diagnosed with Hepatitis C to quantify sodium and potassium content - Analysis of saliva for Calcium, Sodium and Potassium collected from mice modeling dry mouth syndrome. - Analysis of yeast samples and cellular washings for the presence of selenium from incorporated nanoparticles. Sub project #3 ELISA development and automation Automation of Aedes aegypti sera assay for the analysis of IgG in serum samples from people exposed to mosquitos Our unit automated the IgG ELISA developed in Dr Luiz Oliveira's lab and has successfully reproduced the results of the manual assay using automation. Additionally, the assay has been altered to provide for more dynamic range between positive and negative samples. Our unit processed the 4215 samples provided by his lab and additionally processed 921 samples provided by a group at UVA and 199 samples provided by a group at Harvard, which were all provided under material transfer agreements (MTAs) with Dr. Oliveira. The assay will be used to process approximately another 6000 samples provided by a lab at Harvard under an MTA with Dr Oliviera. The assay will also be optimized to determine if other reagents can be used to provide more dynamic range and also investigate IgA and IgM reactivity using the serum samples already provided.