生物化学技术

The polymerase chain reaction (PCR) is a method for the enzymatic amplification of DNA and has revolutionised our approach to both diagnostics and research, permitting the amplification of small amounts of genetic material, either derived from genomic (chromosomal DNA) or RNA via rever ...

Gas chromatography-mass spectrometry (GC-MS) has been used in hormone assays particularly for steroids in biological fluids. The combination of GC with MS exploits the high-resolving power of gas chromatography to separate closely related molecules, and the ability of the MS to provide ...

High-performance liquid chromatography (HPLC) is a refinement of traditional column chromatographic techniques. The speed of analysis and the resolution are increased with new column-packing materials and eluant pumped through the column at high pressure. The potential for ac ...

No other development has had such a major impact on the measurement of hormones as immunoassay. Reagents and assay kits can now be bought commercially but not for the more esoteric or new hormones. This chapter explains the basics of the immunoassay reaction and gives simple methods for immunoass ...

Melatonin and its major metabolite, 6-sulphatoxymelatonin, can routinely be measured by RIA or ELISA. Plasma, serum or saliva samples maybe used for melatonin measurement and urine for the metabolite. Melatonin is a hormone produced by the pineal gland witha clear circadian rhythm giving ...

The gastrointestinal tract is the largest endocrine organ and holds a special place in endocrinology since the concept of blood-borne communication between cells was first established through experiments on the gut. Gut peptide hormones and neurotransmitters regulate the compl ...

Testosterone is the major androgen measured in clinical and research investigations of both men and women. Nevertheless, many other androgens have an important role in t he investigation of andrenal and gonadal physiology and pathology. Commercial assays are generally used in clinic ...

Measurement of angiotensin 1, released in plasma by the action of endogenous enzyme renin on endogenous substrate, angiotensinogen is described. Details are given for the generation of angiotensin 1 from plasma using controlled conditions of pH and temperature. A radioimmunoassay to ...

A detailed method for measuring aldosterone in serum/plasma is described. Aldosterone is first extracted into dichloromethane to improve the specificity of the assay. A radioimmunoassay is performed on the dried extract using an antibody raised in rabbits to aldosterone conjugat ...

General protein export requires the cooperation of two elements, the Sec translocase and a signal sequence. The interactions of both wild type and mutant components can be studied in vivo using a number of genetic systems. Signal sequence mutations that prevent export have been characteri ...

When identical or similar structures exist in different crystallographic environments, similarities between their diffraction patterns, which are directly related to their Fourier transforms, would be expected The technique of Molecular Replacement in protein X-ray crys ...

The determination of the three-dimensional structure of molecules using single crystal X-ray diffraction techniques requires the measurement of amplitudes and the calculation of phases for each diffraction point (maximum). Although amplitudes can be directly measured from d ...

The determination of three-dimensional structures of macromolecules in the crystalline state depends on the acquisition and processing of diffraction data generated by the interaction of X-rays with the crystals of the macromolecule under investigation. In general, there are two ...

During the last 8 years, there has been a revolution in X-ray crystallographic data-collection technology, resulting in an enormous increase in data-acquisition rates and in the range of macromolecules that can be investigated in most laboratories. Provided that the macromolecule fo ...

To determine whether crystals of macromolecules are suitable for crystallographic studies, they must be fully characterized. Characterrzatron is intended to answer many important questions about the crystals. Do they diffract X-rays? Do they diffract to a suitable resolution? Are ...

Rapid developments in recombinant technology have made it possible to overproduce selected proteins of specific interest to the levels required for structural analysis by X-ray crystallography. High-level gene expression has facilitated the purification of many proteins th ...

In recent years, there has been great progress in the determination of high-resolution three-dimensional (3D) structures of membrane proteins. The first major breakthrough came with the crystallization (1) and X-ray crystallography (2,3) of the bacterial photosynthetic reacti ...

Crystallography provides a means of visualizing intact virus particles as well as their isolated constituent proteins and enzymes (1–3) at near-atomic resolution, and is thus an extraordinarily powerful tool in the pursuit of a fuller understanding of the functioning of these simple bi ...

Over the past few years, there has been a dramatic increase in the number of protein-DNA cocrystal structures solved at atomic resolution (Table 1; for reviews see ). This has allowed a detailed understanding of how specific proteins interact with DNA (e.g., prokaryotic repressors), but has not fa ...

Until 1976, the study of nucleic acid structure was exclusively the domain of fiber diffractionists. Between the original Watson-Crick structure in 1953 and this date, there was considerable activity in refining the original B-form model of DNA and extending the approach to other polymor ...