Electrophoresis Protocols

ProtoBlock System

The following reagents are supplied with this system:

  • ProtoBlock Reagent A (100 grams)
  • ProtoBlock Reagent B (200 ml)

To prepare 200 ml of ProtoBlock solution, add 10 grams of ProtoBlock Reagent A to 170 ml of deionized water.  Mix well and add 20 ml of ProtoBlock Reagent B.  This solution is used to block the membrane after protein transfer and to dilute the antibodies used in the immunoassays.

If any blocking solution remains, store at 4oC for future use.  Reconstituted solution is stable at 4-8oC for one week.

ProtoLift Western Stripping Buffer Protocol

Stripping Procedure
 
CAUTION: This procedure requires the use of mercaptoethanol, a hazardous substance. Work in a fume hood.
 
  1. Probe the membrane using your preferred protocol. (do not allow the membrane to dry before stripping).
     
  2. Rinse the membrane in two washes of PBS or TBS. 
     

ProtoStain Blue Protocol

Note: This protocol is optimized for a 0.75mm thick gel. Thicker gels require longer water
washes.
To conserve solution, use a plastic container.
 
  1. Wash gel 3 times at 10 minutes each with deionized water on an orbital shaker. Decant wash solution. Longer washes give better consistency.

  2. After the last wash, add enough ProtoStain Blue solution to completely cover the gel. Invert bottle before use to resuspend particles before dispensing stain solution.

Glass Free Protocol

Glass Free Method of Use
Basic Protocol:
WARNING: Always work with Glass Free in a hood
1. For preparing glass plates for gel casting, Glass Free is to be used only on the removable upper plate. Clean the glass with detergent, water, distilled water, and methanol. 
 
2. After the plate is thorougly dry immerse it in a dish of Glass Free for 5 minutes, making sure that all areas of the glass are well covered by the reagent. Remove any trapped air bubbles. Use latex, PVC or polyethylene gloves.
 

Glass Bond Protocol

Glass Free Method of Use
Basic Protocol

WARNING: Always work with Glass Free in a hood.

UreaGel 29:1 System Protocol

Mix UreaGel System Components:
Determine how much UreaGel 29:1 Concentrate, Diluent, and Buffer you need to make your gels using the formulas below. Combine the necessary components in an Erlenmeyer flask. Swirl gently to mix.

For 100 ml Gel Casting Solution:

Vc = (Vt) (X)/25
Vb = 0.1 (Vt)
Vd = Vt - (Vc + Vb)

SequaGel MD Protocol

Gel Preparation:

  1. Preparing the Working Solutions:
    To cast a 0.8 to 1.0 mm thick gel (>40 cm vertical gel recommended) combine the following in an Erlenmeyer flask:
    • 50 ml SequaGel MD
    • 6 ml 10X TBE
    • 15 g Urea (optional)

    Fill to 100 ml with deionized water and mix thoroughly. Urea may assist the formation of more distinct bands during electrophoresis and reduces the formation of doublets in homoduplex controls.

ProtoGel Quick Cast Protocol

Twenty Minute Casting

ProtoGel Quick-Cast contains the monomers and buffer components to produce a 12% gel.

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