Biology Reference
In-Depth Information
2.3 Equipment
1. Lyophilizer (Labconco).
2. Analytical Balance (Mettler Toledo).
3. Vortex (Velp Scientifi ca).
4. Microcentrifuge (Eppendorf).
5. Ultrasonic homogenizer (Cole Palmer).
6. Spectrophotometer (Eppendorf Bio Photometer).
7. Criterion Stain Free Gel Imaging System (Bio-Rad).
8. Isoelectric Focusing System (PROTEAN IEF cell, Bio-Rad).
9. Electrophoresis systems (PROTEAN II XL, Bio-Rad).
10. FX Pro Plus Multiimager Fluorescence Scanner (Bio-Rad).
11. GS-800 Calibrated Densitometer (Bio-Rad).
12. Quantity One and PDQuest software (Bio-Rad).
2.4 Reagents,
Solutions, and Buffers
Analytical grade reagents must be used, unless other grades are
specifi ed. The prepared solutions should be kept at 4 °C or at
−20 °C if indicated. Reagents and solutions must be discarded
once used, according to current regulations. It is mandatory to use
a fume cupboard when working with volatile and dangerous com-
pounds. Personal protection elements such as robes, gloves and
glasses are of mandatory use.
1. Phenol solution equilibrated with 10 mM Tris-HCl, pH 8
(P4557, Sigma).
2. Bradford reagent (Bio-Rad).
3. ReadyStrip™ IPG strips, 11 cm, pH 3-10 L, and 17 cm,
pH 3-10 L and NL (Bio-Rad).
4. Mineral oil (Bio-Rad).
5. Criterion TGX Stain-Free Precast Gels 4-20 % (Bio-Rad).
6. Precision Plus Protein Standards (Bio-Rad).
7. 30 % Acrylamide/Bis-acrylamide solution (Bio-Rad).
8. SDS-PAGE Broad Range markers 6.5-200 KDa (Bio-Rad).
9. SYPRO Ruby reagent (Bio-Rad).
10. Specialty Standard IEF (Bio-Rad).
11. Total protein extraction buffer: 0.5 M Tris-HCl, pH 8.0, 5 %
(w/v) SDS, 15 % (v/v) glycerol, 100 mM dithiothreitol
(DTT), 1 mM Phenylmethylsulfonyl Fluoride (PMSF).
12. 10 % (w/v) trichloroacetic acid (TCA), 0.07 % DL-dithiothreitol
DTT (w/v) in 80 % (v/v) acetone.
13. 100 % (w/v) trichloroacetic acid (TCA) in water.
14. 0.1 M ammonium acetate in 100 and 80 % (v/v) methanol
solutions.
