Semi quantitative (Western blots) vs Quantitative (ELISA) assays

We often hear the terms quantitative and semi-quantitative in Molecular Biology lab.

Quantitative techniques or assays such as ELISA for determining the amount of protein or real time PCR to measure the amount of cDNA gives us the exact amount or absolute amount of protein or mRNA present within the mixture of other molecules. In order to determine this, standards (various known concentrations of the interested protein) are run in the experiment. This allows us to plot a standard curve which  is used to determine the absolute quantity of the protein. We will be able to say for example that the measured sample has 300pg/ml of a specific cytokine.

Semi-quantitative assay such as Western blots or semi-quantitative RT PCR (not real time) are used to identify and determine the relative quantity of a molecule in a mixture of molecules. This does not give an absolute value which means we wouldn't know how much of sample is present. The only thing we would know is if the sample is present and whether it is more or less (fold change) compared to other samples. We will be able to say for example treated group contains 2 times the protein compared to untreated group.

Do not get confused with the standards used in Western blots. They are run to determine the approximate molecular weight of the proteins not to determine how much protein is present.

OXIDATION & REDUCTION

We generally hear the terms of Oxidation and Reduction in Chemistry and Biochemistry. Students often get confused but there is an easy way to remember these terms.

Oxidation and Reduction are chemical reactions. Always these two reactions occur together. When there is an oxidation, there is always reduction. When one is oxidised the other is reduced.

What is Oxidation & Reduction:

The addition of Oxygen or Removal of hydrogen atoms or Loss of electrons (generally acceptable definition) is Oxidation

The addition of Hydrogen atoms or removal of oxygen atoms or Gain of electrons (generally acceptable definition) is Reduction.

Example: C6H12O6+6O2=6CO2+6H20+ENERGY

In the above chemical reaction, glucose is oxidised (i.e.,12 Hydrogen are removed from the sugar (glucose) molecule) into carbon dioxide whereas oxygen is reduced ( i.e. 12 Hydrogen are added to oxygen atom) into water.
Every Hydrogen atom has one electron, so by removing or adding hydrogen, the same number of electrons are removed or added.

Always remember:

LEO :  Loss of electrons is Oxidation   
GER : Gain of electrons is Reduction