As the saying goes, "Details determine success or failure." Sometimes, small skills in experiments can make a big difference.
Protein Marker: Composed of high-purity proteins with known molecular weights, it serves as a "ruler" to indicate the molecular weight of the target protein, and can also show whether the membrane transfer is successful and the extent of electrophoresis of the protein on the gel. Choosing the right protein Marker is also one of the necessary conditions for the success of Western blot (WB) experiments.
I. Currently, Markers can be roughly divided into three types:
1. Unstained Marker: It does not contain dye molecules or labeling molecules, has no color, and cannot monitor the electrophoresis status and membrane transfer effect of proteins, but it indicates the molecular weight most accurately.
2. Prestained Marker: These contain dye and protein, making them visible to the naked eye. You can directly observe the electrophoresis progress and check if the membrane transfer is complete, which makes them quite commonly used. Each marker band is conjugated with a dye, allowing real-time monitoring of electrophoresis and membrane transfer, as well as reliable localization of the target protein. Note that the molecular weight of pre-stained markers may slightly differ from their actual molecular weight.
3. Developed Protein Marker: These markers contain IgG binding sites that can bind to primary or secondary antibodies used to detect the target protein. This allows the protein marker to appear on the membrane alongside the target protein band. Usually, two pre-stained bands are included for real-time observation of electrophoresis and membrane transfer progress, and to assess transfer efficiency and direction.
II. How to Choose a Protein Marker?
Firstly, clarify the experimental purpose. If you only need to observe the size of protein bands after electrophoresis, obtain accurate protein molecular weights, or distinguish between closely sized bands, an unstained protein Marker can be used. If performing a WB experiment, a prestained protein Marker can be chosen to observe the electrophoresis and membrane transfer in real-time.
Here are some factors to consider when selecting a suitable protein marker:
1. Molecular Weight Range: Protein markers are categorized into high, low, and broad molecular weight ranges. High molecular weight markers are commonly used for detecting large proteins, while low molecular weight markers are used for smaller proteins or peptides. If you need flexibility across your lab, choosing a broad molecular weight marker with evenly distributed bands can help you identify proteins in any molecular weight range.
2. Band Accuracy: Servicebio high-purity recombinant protein markers are ideal for use as molecular weight standards in SDS-PAGE and Western blot applications.
3. Quality and Stability: Servicebio markers undergo multiple washings and long incubation times without band loss or fading. They have high affinity for the membrane, ensuring stable band color and reliable results.
