生物化学技术

Human plasma is regarded the most complex and well-known clinical specimen that can be easily obtained; alterations in the levels of plasma proteins or their corresponding enzyme activities may reflect either a healthy or a diseased state. Given that there is no defined genomic information ...

Laser capture microdissection (LCM) is a powerful tool for procuring near-pure populations of targeted cell types from specific microscopic regions of tissue sections, by overcoming problems due to tissue heterogeneity and minimizing intermixture and contamination by other c ...

Difference gel electrophoresis (DIGE) technology has been used to provide a powerful quantitative component to proteomics experiments involving 2D gel electrophoresis. DIGE combines spectrally resolvable fluorescent dyes (Cy2, Cy3, and Cy5) with sample multiplexing for low ...

The ability to visualize the full depth of the serum proteome in a high-throughput manner is a major goal of clinical proteomics. Methodologies, which combine higher throughput with the ability to observe differential protein expression levels, have been applied to this goal. An example of s ...

Urine represents the most easily attainable and consequently one of the most common samples in clinical analysis and diagnostics. However, urine is also considered one of the most difficult proteomic samples to work with due to its highly variable contents, as well as the presence of various pr ...

Laser microdissection is an effective technique to harvest pure cell populations from complex tissue sections. In addition to using the microdissected cells in several DNA and RNA studies, it has been shown that the small number of cells obtained by this technique can also be used for proteomics ...

Isotope-coded affinity tag (ICAT) labeling, in combination with mass spectrometry (MS), has been widely adopted as an effective method for comparing protein abundance levels. This chapter describes the ICAT labeling procedure in search for the celecoxib-regulated proteins in a col ...

Laser capture microdissection (LCM) is a powerful tool that enables the isolation of specific cell types from tissue sections, overcoming the problem of tissue heterogeneity and contamination. We combined the LCM with isotope-coded affinity tag (ICAT) technology and two-dimensio ...

Pharmaceutical companies and regulatory agencies are pursuing biomarkers as a means to increase the productivity of drug development. Quantifying differential levels of proteins from complex biological samples like plasma or cerebrospinal fluid is one specific approach be ...

The extracellular matrix (ECM) and secreted vesicles are unique structures outside of cells that carry out dynamic biological functions. ECM is created by most cell types and is responsible for the three-dimensional structure of the tissue or organ in which they are originated. Many cells al ...

This chapter describes the development and use of bead-based miniaturized multiplexed sandwich immunoassays for focused protein profiling. Bead-based protein arrays or suspension microarrays allow simultaneous analysis of a variety of parameters within a single experim ...

Antibody arrays represent one of the high-throughput techniques enabling detection of multiple proteins simultaneously. One of the main advantages of the technology over other proteomic approaches resides on that the identities of the measured proteins are known at front of the expe ...

Due to the low reproducibility affecting 2D gel-electrophoresis and the complex maps provided by this technique, the use of effective and robust methods for the comparison and classification of 2D maps is a fundamental tool for the development of automated diagnostic methods. A review of cl ...

After separation through two-dimensional gel electrophoresis (2DE), several hundreds of individual protein abundances can be quantified in a cell population or sample tissue. Both a good experimental setup and a valid statistical approach are essential to get insight into the data and ...

Current proteomics technologies generate large number of data among which the investigator has to identify the promising diagnostic/prognostic biomarkers as well as potential therapeutic targets. For the latter, classification of proteins into meaningful families is need ...

The analysis of protein mixtures by liquid chromatography-mass spectrometry (LC-MS) requires tools for viewing and navigating LC-MS data, locating peptides in LC-MS data, and eliminating low-quality peptides. msInspect, an open source platform, can carry out these steps for single e ...

The statistical classification strategy we have developed for magnetic resonance, infrared, and Raman spectra for the analysis of biomedical data is discussed, particularly as it applies to proteomic mass spectra. A general discussion of the current use of pattern recognition metho ...

A comprehensive collection of different methods for extracting high-quality genomic DNA from Gram-positive and -negative bacteria and fungal mycelium and spores is described in this chapter. Special care has been taken in describing the ideal ratio of biological material to chemic ...

A misconception regarding the sensitivity of nonradioactive methods for screening genomic DNA libraries often hinders the establishment of these environmentally friendly techniques in molecular biology laboratories. Nonradioactive probes, properly prepared and qu ...

Several specific problems are encountered when nonradioactive detection methods are used in conjunction with plant nucleic acids. In this chapter, we describe protocols for the isolation of DNA and RNA from plant leaves and the preparation of probe molecules by either PCR or in vitro transc ...