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Bio-Rad ReadyPrep Sequential Extraction Kit User Manual

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Sequential extraction can be carried out in a single

microcentrifuge tube, minimizing protein loss. The kit is

particularly well suited for use with 2-D PAGE systems

employing immobilized pH gradients for the first-dimension

IEF.

Section 2. Kit Components

Reagent 1. One vial. Lyophilized. Each vial of rehydrated

Reagent 1 contains 50 ml of 40 mM Tris base.

Reagent 2. Three vials. Lyophilized. Each vial of rehydrated

Reagent 2 contains 10 ml of 8 M urea, 4% (w/v) CHAPS, 40

mM Tris, and 0.2% (w/v) Bio-Lyte 3/10 ampholyte.
Reagent 3. Two vials. Lyophilized. Each vial of rehydrated

Reagent 3 contains 10 ml of 5 M urea, 2 M thiourea, 2% (w/v)

CHAPS, 2% (w/v) SB 3-10, 40 mM Tris, and 0.2% (w/v) Bio-

Lyte 3/10 ampholyte.
Reducing Agent TBP. One ampoule containing 0.6 ml of

200 mM tributyl phosphine (TBP) in 1-methyl-2-pyrrolidinone

(NMP) sealed under nitrogen gas.
Empty Vial. One storage vial for reducing agent TBP.

Instruction Manual.

Note: CHAPS is 3-[(3-Cholamidopropyl)dimethylammonio]-1-

propanesulfonate, a zwitterionic detergent.
SB 3-10 is N-decyl-N,N-dimethyl-3-ammonio-1-

propanesulfonate, or caprylyl sulfobetaine, a zwitterionic

detergent.
Bio-Lyte 3/10 is a mixture of carrier ampholytes, pH 3–10.

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