Biochemical Techniques

Biochemical techniques involve the use of chemicals, enzymes, and other biological molecules to analyze and manipulate biological systems. These techniques are used to detect, quantify, and analyze biochemical and molecular processes.

Biochemical analysis techniques

Biochemical analyses are an array of methods, assays, or procedures that allow researchers to study the substances that occur on living bodies as well as the underlying chemical reactions.

The most sophisticated techniques are reserved in specialty research laboratories although simple sets of techniques are employed in common events including testing illicit drugs in competitive sports and analyzing blood sugar in diabetic athletes.

Basic Techniques

Common techniques include immunoassays, chromatography, spectroscopy, and PCR. These techniques can be used to study proteins, DNA, RNA, and other biological molecules.

Chromatography

Chromatography is a technique used to separate components of a mixture based on their interactions with a stationary phase and a mobile phase.

Spectroscopy

Spectroscopy is a technique used to measure the amount of energy absorbed or emitted by molecules during a chemical reaction. It is used to identify the chemical composition of a sample and to measure the concentrations of different molecules.

Electrophoresis or Gel Electrophoresis

Electrophoresis is a technique used to separate charged molecules in a sample based on their size and charge. It is commonly used to separate proteins and nucleic acids.

In this process, molecules are placed in a gel medium and an electric current is applied to them. The molecules move through the gel in response to the electric field, and the different molecules are separated based on their size and charge.

Enzymatic Assays

Enzymatic assays are used to measure the concentrations of enzymes in a sample. It is also used to measure the activity of enzymes in a sample.

Immunoassays

Immunoassays are used to detect the presence of specific molecules in a sample. It is commonly used to measure the concentration of hormones and other proteins in a sample.

Western Blotting

Western blotting is a technique used to separate and identify proteins in a sample. It is commonly used to detect the presence of specific proteins in a sample.

Centrifugation

Centrifugation is a process used to separate particles from a mixture based on their size, shape, and density. It is used in a variety of applications, including medical diagnostics and laboratory studies. The mixture is placed into a centrifuge and spun at high speeds in order to separate the heavier particles from the lighter particles.

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Immunoprecipitation Assays

Immunoprecipitation assays are a type of laboratory technique used to study protein-protein interactions. In these assays, antibodies specific to a particular target protein are used to isolate that protein from a complex sample.

The antibody-protein complex is then isolated from the sample and the protein of interest can be further analyzed. This type of assay is useful for studying the function and interaction of proteins in various biological systems.

It can be used to identify new protein-protein interactions, elucidate protein pathways and networks, and study the effects of drugs or other compounds on protein activity.

Fluorescence Techniques

Fluorescence techniques are used in a variety of scientific and medical applications. In general, fluorescence is a process that occurs when the light of a certain wavelength is shone onto an object and it is then re-emitted at a different wavelength. This is usually done by using a light source such as a laser or a lamp and then detecting the light emitted from the object.

The light that is re-emitted can be used to detect the presence of certain molecules or materials, to measure the concentration of certain molecules, or even to detect the presence of specific cells or other biological components. One of the most popular fluorescence techniques is fluorescence spectroscopy.

This technique is used to measure the concentration of particular molecules in a sample. By shining a light source on a sample, the fluorescence emitted from the sample can be measured and compared to the light source. This technique is used to measure the concentration of molecules, such as proteins and enzymes, in a sample.

Radioimmunoassay (RIA)

It is a laboratory technique used to measure the level of a specific substance, such as a hormone or drug, in a sample of body fluid. RIA works by using radioactively labeled molecules that are attracted to the specific substance in the sample, allowing the substance to be quantified.

RIA is a very sensitive technique, capable of detecting substances present in very small amounts. It is commonly used in medical diagnosis, as well as in drug testing.

Spectrophotometry

It is a technique used to measure the intensity of light as a function of its color, or wavelength. It is used in a variety of applications, including the measurement of the concentration of a sample in a solution, the identification of unknown compounds, and the determination of the purity of a sample.

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In a spectrophotometer, light passes through a sample, and the intensity of the light transmitted is measured with a detector. From the intensity of the light, the concentration of the sample can be determined. This technique is widely used in the fields of chemistry and biology.

Thin Layer Chromatography (TLC)

It is a chromatographic technique used to separate and identify components of a mixture. It involves the use of a thin layer of adsorbent material, such as silica gel or alumina, coated onto a plate or sheet.

Separation occurs when the components of the mixture are distributed between the mobile phase (solvent) and the stationary phase (adsorbent material). As the mobile phase moves up the plate, the components of the mixture move along with it and interact differently with the stationary phase, resulting in different rates of migration.

By plotting the distance traveled by each component of the mixture, a chromatogram can be created which can be used to identify the components of the mixture. TLC is a useful tool in the analysis of pharmaceuticals, food products, and environmental samples.

Polymerase Chain Reaction (PCR)

PCR is used in molecular biology for amplifying and copying a specific piece of DNA. It is used in research, diagnostics, and forensics to detect and measure the presence of a particular gene or nucleotide sequence in a sample.

PCR can be used to amplify DNA from a single cell or from a sample of many cells. The process involves heating and cooling the sample to separate the two strands of DNA, before adding an enzyme called a polymerase and specific nucleotides.

These nucleotides rebuild the two strands of DNA, with each cycle of the process doubling the number of copies of the target DNA sequence. PCR is a fast, simple, and reliable technique that is used to obtain millions of copies of a specific DNA sequence within a few hours.

Tissue culture

It is the process in which pieces of tissues or isolated cells are cultivated in a special fluid medium in a container of glass, plastic, tubes, vials, or bottles.

Chromatography

It is another important biochemical technique to separate small qualities of organic (and inorganic) compounds depending upon the rate of their flow on the chromatographic paper.

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X-ray Crystallography

It is used to determine the molecular organization of substances. The molecular configuration of a double helix of DNA was determined by Watson and Crick through the application of this biochemical technique.

Enzyme-Linked Immunosorbent Assay (ELISA)

ELISA is a laboratory technique used to detect and measure antibodies and antigens in a sample. It is commonly used for medical diagnosis and research, such as testing for HIV or measuring levels of hormones in the blood.

The assay uses an enzyme-labeled antigen or antibody to bind to a target analyte in the sample, resulting in a detectable signal. ELISAs offer advantages such as accuracy, sensitivity, and the ability to measure multiple analytes in one sample.

Methods in biochemistry

Biochemistry methods involve a wide variety of techniques used to identify, isolate, and analyze macromolecules such as proteins, carbohydrates, lipids, and nucleic acids.

Some of the common methods used in biochemistry are chromatography, electrophoresis, spectroscopy, X-ray crystallography, mass spectrometry, nuclear magnetic resonance, and enzyme-linked immunosorbent assays.

Each technique has its own advantages and disadvantages, and choosing the appropriate method for a given project depends on the research goals and the available resources.

In addition, some methods are used to prepare samples for use in other techniques, such as cell fractionation and protein purification.

Errors

  1. Incorrect sampling: If a sample is not taken from the correct location, the analysis results will be inaccurate.
  2. Incorrect storage: If a sample is not stored properly, its composition can change and the analysis results will be inaccurate.
  3. Improper handling: If the sample is not handled correctly during collection and analysis, the results will be inaccurate.
  4. Contamination: If the sample is contaminated with foreign substances, the analysis results will be inaccurate.
  5. Improper calibration: If the instruments used to measure the sample are not properly calibrated, the results will be inaccurate.
  6. Operator error: If the operator performing the analysis makes a mistake, the results will be inaccurate.
  7. pH error: For biochemical analysis, the pH of a solution must be accurately measured by a pH meter otherwise the may be inaccurate.

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