Immuno Dtection

SIITUB

The “SIITUB” is intended as a reference to briefly present the experimental methods frequently used in biochemistry. These are not experimental protocols as such. For each of the methods, it presents the basic principles, methods and devices, the main applications to biochemistry, etc. The SIITUB is under “permanent construction”.

New sections will be added as we go. Corrections and updates will be made, if necessary, to the texts already displayed. The date of the last version is indicated in brackets following the description of the content of each section. The same section can appear in several headings if this is relevant.

BASIC LABORATORY METHODS: SOLUTIONS AND TA
pH and buffered solutions: preparation of buffered solutions, comments on some commonly used buffer products (980504)
Physiological solutions: use and description of certain physiological solutions, preparation, sterilization (04 06 14)

GENERAL METHODS
Homogenization: methods and devices for homogenizing tissues and cells (980127)
Dialysis: principles, description, various applications, concentration calculations (07 10 03)
Lyophilization: principles, equipment, basic techniques (980716)
Filtration and ultrafiltration: types of filter, filtration devices, ultrafiltration, micro-concentration, cold sterilization (950504)
Detergents and protein denaturation: various types of detergents and chaotropic agents (00 02 23)
DOSAGES
Enzyme assay: enzyme and rate units, techniques for measuring activities and rates, derivation of kinetic parameters, etc. (00 01 07)
Protein Assay: Principles, Advantages and Weaknesses of the Main Protein Assay Methods (980407)

CHROMATOGRAPHY
Gel filtration: principles, main gels, dead volume and column calibration, desalting, molecular mass determination, protein purification, columns, samples, elution (99 05 10)
Affinity chromatography: principles, description of the main steps, examples of protein and nucleic acid chromatography (98 05 05)

ELECTROPHORESIS OF PROTEINS
Electrophoresis: general principles of electrophoresis, brief presentation of the most common types of electrophoresis (polyacrylamide gel, agarose, cellulose acetate, etc.) (03 02 00)
Acrylamide gel protein electrophoresis: principles, polymerization, EGPA-SDS, mass estimation (12 05 03)
Isoelectric focusing (FIE) and two-dimensional electrophoresis: principles of protein separation according to pI, creation of mobile and immobilized gradients, two-dimensional electrophoresis
Blotting (Electrophoretic transfer of proteins – Western blot): general description of the methods of electrphoretic transfer of proteins and direct application of proteins, transfer conditions, immunodetection

PROTEINS and ENZYMES
Protein precipitation: basic principles, precipitation with solvents, with acids, with salts, differential precipitation, importance in purification (04 03 03)
Protein assay: principles, advantages and weaknesses of the main methods of protein assay and determination of kinetic parameters
Protein purification: purification strategies, conservation and storage, purification table
Detergents and protein denaturation: various types of detergents and chaotropic agents
Enzyme assay: enzyme and rate units, techniques for measuring activities and rates, derivation of kinetic parameters, etc. (24 02 04)
Methods of studying LDH: determination, separation of subunits, etc. (08 01 03)
Concanavalin A: structure, properties, applications
CENTRIFUGATION
Centrifugation: general principles, equations, centrifuges, rotors, speed calculation …
Gradient centrifugation: discontinuous and continuous gradients, preparation of gradients, rotors, zonal and isopycnic separations …

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