LDH Lab 4 Protocol

LDH Lab 4 Protocol - Section N TA: Josh LDH Lab 4 Protocol...

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Section N TA: Josh LDH Lab 4 Protocol Procedure: 1) Use SDS-PAGE to determine the subunit Mr or LDH. 2) Use gloves when working with acrylamide. 3) Make sure equipment is clean before assembling gel apparatus. 4) Lay longer glass plate down, then place 2 spacers along short edges of the plate. 5) Place shorter glass plate on top of spacers to align (spacers should be sticking up >5 mm above the long glass plate). 6) Loosen 4 screws on clamp assembly and position it so the screws point away from you. 7) Gently slide plate into clamp assembly with longer plate facing away (see figure E2 in lab manual). 8) Tighten the screws. 9) Place assembly into alignment slot on casting stand so screws face away. Loosen the top 2 screws to allow plates and spacers to settle against the casting stand base. Spacers should be vertically flush with sides of sandwich clamp. Tighten both pairs of screws. *Make sure spacers and plates are aligned and everything flush. 10) Pull sandwich from alignment slot and make sure plates and spacers are flush at the bottom. 11) Put casting stand on level ground, check to see gaskets are clean and place on top of red foam pads of the casting stand slots. 12) Transfer clamp assembly to one of the casting slots in the stand. 13) To attach sandwich: butt acrylic pressure plate against wall of casting slot at bottom so that the plates rest on the gasket. *Plates and spacers should stick out slightly. 14) Snap acrylic pressure plate underneath the overhang of casting slot by pushing white portions of clamp. *Do not push against plates or spacers. 15) Place comb between glass plates so that edges sit on spacers. Use sharpie to mark glass 1 cm below the bottom of the comb. This is the max fill line for the resolving gel. Take comb out. 16) Add distilled water to fill line with water bottle or pipet.
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This note was uploaded on 05/10/2008 for the course BIO CHEM 153L taught by Professor Kim during the Spring '08 term at UCLA.

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LDH Lab 4 Protocol - Section N TA: Josh LDH Lab 4 Protocol...

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