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High-Throughput Clinical Proteomics Core | HTCP
The High-Throughput Clinical Proteomics (HTCP) core laboratory is a mass spectrometry core facility designed to profile large numbers of clinical samples for changes in protein content. We use small amounts (micrograms) of protein from samples such as urine, plasma, CSF, and cell lysates and incubate them with magnetic beads that bind different subsets of the total proteome. We then examine the different subsets of small proteins and peptides by high-resolution MALDI-TOF mass spectrometry. We measure the amounts and intact masses of the proteins in the samples and determine which have the greatest statistical relevance in defining clinical groupings before moving to physically isolate those peptides/proteins and identify them. The method works best for peptides and small proteins below 25 kDa and is an excellent complement to gel-based methods such as DIGE.
Goals & Mission.
The HTCP core laboratory was established specifically to assist those engaged in clinical research at UCLA in the analysis of protein profiles of large number of patient samples. We are set up to use robotic methods for the extraction of intact proteins and peptides from clinical isolates followed by analyses of protein patterns that provide the most diagnostic information on groupings in the patient population. We do not identify as many proteins as possible in these samples; we isolate and ID only those determined to be of clinical significance. Our approach is to validate these biomarker candidates in a relevant number of patient samples (dozens to hundreds) before identification. While we usually study intact masses of proteins and peptides, we can also analyze those subjected to protease digestion.
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Services
Protein Profiling: The main service that the core facility provides is high-throughput protein profiling using solid-phase extraction of proteins/peptides with magnetic beads followed by high-resolution MALDI-TOF mass spectrometry. We develop methods to maximize the number of detected proteins specific to particular types of samples such as plasma, urine, saliva, CSF, and cell extracts. We analyze the large number of sample mass spectra to determine those protein signals of greatest significance and can isolate those proteins by HPLC. Profiling data can also be provided directly to the collaborating lab for analysis using a variety of approaches such as microarray formats.
Protein Isolation and Identification: Biomarker candidates are physically isolated from the clinical samples using a variety of techniques such as ultrafiltration, ion-exchange fractionation, and semi-preparative and capillary HPLC. Proteins are typically digested with trypsin if greater than 4-5 kDa and the peptides are fragmented using a variety of mass spec techniques such as LC-MALDI MS/MS and electrospray MS/MS.
Tissue Imaging: The core recently took delivery of a new MALDI TOF-TOF instrument (Bruker UltrafleXtreme) which is capable of detecting proteins directly from thin tissue sections mounted to conductive glass microscope slides. We will be capable of compiling mass spectra across the tissue with 25-200 um resolution and converting them into heat maps based on protein or lipid abundance. This service is still being developed and should be available in late 2011/early 2012.
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Ordering
Typical charges for protein profiling analyses of clinical samples are $35-135 each (run in triplicate) depending on the complexity of the samples and whether extensive prefractionation and depletions of abundant plasma proteins are required. MALDI Tissue imaging typically costs from $200 to $500, depending on the size of the frozen section and which resolution is required. Protein isolation and ID charges are made for each individual step required.
Please contact us for more information.
Phone: (310) 825-8753 | Email
