Question

In chemistry, define the difference between an absolute standard versus a relative standard, and give examples. Also, what factors affect the choice of standard protein used.

Answer 1

Factors that affect the choice of standard protein used

There are various factors involved in affecting the choice of standard protein used, In addition to the nature of the protein. They are as follows:

Preparation of the Sample

Sample Preparing method may be the factor. For that Solubilisation of Protein Samples in a non interfering agents with buffer or interfering agents should be eliminated by dialysis before analysing.

Interfering agents

While handling with interfering agents, using copper ions for estimation of proteins which contains metal cheating agents, dyes, reducing agents, sugars and amines is restricted when dye based proteins is not able to used for protein solutions which has detergents or surfactants. While doing this it results in a false values. For attaining exact protein estimation the choice of assays there should not be a non protein agents or it could be removed considerably from the protein solutions. The alternative solution is to follow the procedure that avoid the interfering actions in the protein samples by non protein agents.

Sample Size and Sensitivity

When the sample is insufficient in availablity, the size of the sample for estimation of protein plays a crucial role. In that time, better go for the method which requires less amount of protein sample for accurate estimation.

Protein Sample in diluted state

Usually 0.5$\mu$g proteins required for accurate estimation calorimetric protein assays , whereas for achieving the detection limit dilute protein consumes larger volume. When the amount of sample is 10% larger than the total assay will leads to the interference of volume in most of the assays. By choosing one which focus the samples as normal course of assay method. This prevent the results from negatively affected by dilute samples.

Assay time and Consideration of time

Determination of the time needed to achieve a protein assay is carried out by assay method and the sample complexity. Samples with interfering agents and Standard plots or curves may consume more time, but using protein assays which does not depend upon standard plots for protein estimation, with less samples.

Standard of Proteins

It is a tedious process by using a standard, similar to the sample for the execution of protein estimation. It is relaxed by using readymade proteins(gamma globulin and BSA) as standards. This in result no dependency for choosing protein standards exhibited by the standards.

Variation in Protein to Protein

Protein to Protein Variation is mainly viewed in Dye based Proteins. There may be circtumstances of no protein reponse, because of the absence of the protein dye complex.

Instrumentation Specification

For high throughput applications Multi well titer plates are used. In general colorimeters and spectrophotometers are demanded by most of the protein assays. Regarding titer plates not all the protein assays are able to carried out.

Difference between absolute standard and relative standard

Absolute Standard values are the exact values which can be used as a reference for experiments, whereas relative standard values the approximate values, the results may be the nearby values which can be increased or decreased.

Obviously Absolute Standard is selected, Because the values are fixed. But in the relative standard there are some disadvantages. The convenience is relative standard is it gives way for correcting the change in final crisis.

Pure Samples are Standard samples It shows various types of similarity to the experiment sample. The relative standard is also absolute standard divided by some other standard.