Medical research

Biomedical research (or experimental medicine), in general simply known as medical research, is the basic research, applied research, or translational research conducted to aid and support the body of knowledge in the field of medicine. Medical research can be divided into two general categories: the evaluation of new treatments for both safety and efficacy in what are termed clinical trials, and all other research that contributes to the development of new treatments. The latter is termed preclinical research if its goal is specifically to elaborate knowledge for the development of new therapeutic strategies. A new paradigm to biomedical research is being termed translational research, which focuses on iterative feedback loops between the basic and clinical research domains to accelerate knowledge translation from the bedside to the bench, and back again.

Protein-DNA Interactions

• ChIP-on-chip
• Chip-Sequencing
• DamID

Protein structures

• X-ray crystallography
• Protein NMR

Protein purification

• Protein Isolation
  • chromatography methods
• Protein Extraction and Solubilization
• Protein Concentration Determination Methods, Bradford protein assay
• Concentrating Protein Solutions
• Gel electrophoresis
  • Gel Electrophoresis Under denaturing conditions
  • Gel Electrophoresis Under non-denaturing conditions
  • 2D Gel Electrophoresis
• Electrofocusing

Detecting proteins

• microscopy, Protein immunostaining
• Protein immunoprecipitation
• Immunoelectrophoresis
• Immunoblotting
• BCA Protein Assay
• Western blot
• Spectrophotometry
• Enzyme assay

Genetic methods

• conceptual translation- many proteins are never directly sequenced, but their sequence of amino acids is known by "conceptual translation" of a known mRNA sequence. See Genetic code.
• site-directed mutagenesis allows new variants of proteins to be produced and tested for how structural changes alter protein function.
  • insertion of protein tags such as the His-tag. See also: Green fluorescent protein.
• evolutionary; analysis of sequence changes in different species using software such as BLAST.
• Proteins that are involved in human diseases can be identified by matching alleles to disease and other phenotypes using methods such as calculation of LOD scores.

Protein methods

Protein methods are the techniques used to study proteins.

There are genetic methods for studying proteins, methods for detecting proteins, methods for isolating and purifying proteins and other methods for characterizing the structure and function of proteins, often requiring that the protein first be purified.

Macromolecule blotting and probing

The terms northern, western and eastern blotting are derived from what initially was a molecular biology joke that played on the term Southern blotting, after the technique described by Edwin Southern for the hybridisation of blotted DNA. Patricia Thomas, developer of the RNA blot which then became known as the northern blot actually didn't use the term. Further combinations of these techniques produced such terms as southwesterns (protein-DNA hybridizations), northwesterns (to detect protein-RNA interactions) and farwesterns (protein-protein interactions), all of which are presently found in the literature.

Polymerase chain reaction

The polymerase chain reaction is an extremely versatile technique for copying DNA. In brief, PCR allows a single DNA sequence to be copied (millions of times), or altered in predetermined ways. For example, PCR can be used to introduce restriction enzyme sites, or to mutate (change) particular bases of DNA, the latter is a method referred to as "Quick change". PCR can also be used to determine whether a particular DNA fragment is found in a cDNA library. PCR has many variations, like reverse transcription PCR (RT-PCR) for amplification of RNA, and, more recently, real-time PCR (QPCR) which allow for quantitative measurement of DNA or RNA molecules.

Expression cloning

One of the most basic techniques of molecular biology to study protein function is expression cloning. In this technique, DNA coding for a protein of interest is cloned (using PCR and/or restriction enzymes) into a plasmid (known as an expression vector). This plasmid may have special promoter elements to drive production of the protein of interest, and may also have antibiotic resistance markers to help follow the plasmid.

This plasmid can be inserted into either bacterial or animal cells. Introducing DNA into bacterial cells can be done by transformation (via uptake of naked DNA), conjugation (via cell-cell contact) or by transduction (via viral vector). Introducing DNA into eukaryotic cells, such as animal cells, by physical or chemical means is called transfection. Several different transfection techniques are available, such as calcium phosphate transfection, electroporation, microinjection and liposome transfection. DNA can also be introduced into eukaryotic cells using viruses or bacteria as carriers, the latter is sometimes called bactofection and in particular uses Agrobacterium tumefaciens. The plasmid may be integrated into the genome, resulting in a stable transfection, or may remain independent of the genome, called transient transfection.