Simon's Protocols

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Suppliers:

General Lab Supplies:

VWR

Antibodies:

Cell Signaling

Covance

EMD Biosciences (Calbiochem)

R & D Systems

Santa Cruz

Dissection Tools:

FST

WPI

Misc:

Assay Design

Bangs Labs (Silica beads)

Biorad

Biolynx

BD-Biosciences

Carolina (coverslips)

Chemicon

Cytoskeleton (G-LISA kit)

Invitrogen

MatTek (glass bottom dishes)

MP Biomedicals (formely ICN Biochemicals)

Promega

General:

Recipes:
o Buffers
o Drug, protease & phosphatase inhibitor dilutions

Cell culture:
o Media comparison (DMEM, Hanks', L15, NeuroB, B27 and N2)
o Substrates ***
o Bacteria
o General HEK 293T culture ***

Primary cells:
o Rat E12 spinal cord ***
o Rat E14 dissociated commissural neurons *** (videos)
o Rat E14 dorsal explant *** (videos)
o Rat E15 DRG (dissociated and explants)** (video)
o Corticals **

Protein methods:
o SDS-PAGE ***
o Staining & drying acrylamide gels ***
o Western blot (with ponceau and india ink stains) ***
o Immunoprecipitation **
o Direct fluorescent labeling *
o Netrin, Slit Purification ***

Nucleotide methods:
o Agarose Gel ***
o Maxiprep ***
o RNA Isolation (using RNeasy)
o RT-PCR (using Qiagen's One step kit)

Immunofluorescence:
o Dissociated cells ***
o Embryos*
o Explants ***

Specialized:

cAMP & pCREB:
o Phospho-CREB analysis ***
o cAMP ELISA ***

Rho GTPase:
o GST-RBD purification ***
o RhoA pulldown with GST-RBD ***
o RhoA in PM *
o G-LISA ***

Adhesion:
o Cellular adhesion ***
o Immobilized circular protein pattern on a coverslip **

Plasma Membrane Localization:
o Biotinylation *** (just PM proteins)
o Biotinylation *** (PM and endocytosed proteins)
o Immuno-labeling of proteins exposed to cell surface **

HSV Gene Transfer:
o Plasmid Purification *
o Virus Purification *

Microscope:
o DIC
o Replace & adjust fluorescent bulb **
o Confocal

Laser Trap:

o Calibration ***

o Bead prep ***

o Laser trap ***

Pillars:

o Polyacrylamide Pillars ***

Online resources:

Biocompare (great for findings reagents, kits, antibodies...)

Cell & Molecular Biology Online

Current Protocols (excellent... but requires a subscription)

Laboratory Protocol List (a huge list of biochemistry protocols)

MGAlign (determines exons from mRNA & genomic sequences)

Protocol online (a well organized, general protocol database)

Primer 3 (great for designing PCR primers)

Books:

Culturing Nerve Cells

Protocols for Neural Cell Culture

Stars (*) indicate confidence level:
* = in development (low)
** = works, but not extensively tested
*** = Used often

Last update: 05/31/2009

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