Biology Reference
In-Depth Information
HBSS: Hank's balanced salt solution (can be prepared as 5
buffer and stored at
4
C); 1
solution can be obtained from Gibco (#14025-050)
1.26 mM CaCl
2
5.33 mM KCl
0.44 mM KH
2
PO
4
0.5 mM MgCl
2
0.41 mM MgSO
4
138 mM NaCl
4 mM NaHCO
3
0.3 mM Na
2
HPO
4
5.6 mM glucose
MTSB: Microtubule stabilizing buffer (store at 4
C)
1 mM EGTA
4% PEG 8000
100 mM PIPES, pH 6.9
TSB
0.5% Triton X-100 in MTSB
PFA in MTSB
4% PFA in MTSB: PFA powder (Sigma #P6148) is heated in MTSB to
approximately 50
C and dissolved by adding a drop of 10 M NaOH; the
pH is then brought to pH 7.0 with 6 M HCl. The solution is left to cool
down to RT.
Mowiol mounting medium
10% Mowiol (polyvinyl alcohol) 4-88 (Sigma #81381)
100 mM Tris/HCl, pH 8.5
25% glycerol
16.3.2
Gel electrophoresis and immunoblotting
16.3.2.1
SDS-PAGE for
a
/
b
-tubulin separation
Laemmli sample buffer
450 mM DTT
10% SDS (BDH #442444H)
400 mM Tris/HCl, pH 6.8
50% glycerol
bromophenol blue
Electrophoresis cell: Mini-PROTEAN
®
cell with a PowerPac
™
power supply
(Bio-Rad #165-8001EDU and #164-5052MP, respectively).
5
Separating gel buffer (4
)
1.5 M Tris/HCl, pH 9.0
0.4% SDS (specific type required: Sigma #L5750)
