[Histonet] VEGF and VEGF-R in FFPE mouse

[Mack, Sarah]

I hope that this will help.  Best of luck. 

VEGF  Immuno Protocol 						
Date Run:			
Step						Minutes		Temp
Complete
1) Bake slides  				60 			60C	
2) Deparaffinize & rehydrate to PBS				RT	
3) Methanolic 3% H2O2  			30			RT	
4) Rinse PBS X 3 				15			RT

5) DAKO Proteinase K Digestion	6			RT	
6) Rinse PBS X 3				5			RT	
7) PAP around section						RT	
8) 10% Normal Goat Serum/PBS/BSA	30			RT	
9) VEGF 1:200/PBS/BSA			Overnight		4C	
10) Rinse PBS X 3				15			RT

11) Bo Goat anti-rabbit IgG 1:500	30			RT	
12) Make ABC 							RT	
13) Rinse PBS X 3				15			RT

14) ABC complex				30			RT	
15) Rinse PBS X 3				15			RT

16) 0.5% DAB				3-10 min		RT	
17) Rinse DH2O				3			RT	
18) Counterstain Hematoxylin		1			RT	
19) Dehydrate to Xylene						RT	
20) Cover with Permount					RT	
			
Reagent Lot Numbers:			
Normal Goat Serum : Vector S-1000 : Lot #			
VEGF  Antibody : Santa Cruz SC-507 : Lot #			
Bo  Goat  anti-rabbit : Vector BA-1000 : Lot #			
ABC Kit : Vector PK:6100 : Exp. 			
DAB Kit : Vector SK:4100 : Exp.			
			
Notes:			
                 1)All antibodies made with PBS  	 &1% BSA & 0.1%
Tween		
                 2)Negative control Rabbit IgG 1:200			
                 3) DAKO Digestion:1 Drop stock/ 	 2ml .05M Tris 	HCl
pH7.5	

> ----------
> From: 	Trajkovic, Dusko
> Sent: 	Wednesday, August 24, 2005 9:23 AM
> To: 	Mack, Sarah
> Subject: 	RE: [Histonet] VEGF and VEGF-R in FFPE mouse
> 
> Good morning Sarah,
> I think many of us would love to see a complete protocol procedure for the
> VEGF antibody. In the past, I have tried many different procedures and
> none
> of them resulted in any meaningful success.
> Thank you in advance and have a wonderful day.
> Dusko Trajkovic
> 
> -----Original Message-----
> From: histonet-bounces <@t> lists.utsouthwestern.edu
> [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Mack,
> Sarah
> Sent: Wednesday, August 24, 2005 5:43 AM
> To: histonet <@t> lists.utsouthwestern.edu;
> 'wasielewski.reinhard.von <@t> mh-hannover.de'
> Subject: RE: [Histonet] VEGF and VEGF-R in FFPE mouse
> 
> We use Santa Cruz VEGF antibody, catalog #SC-507.
> 
> 
> Sarah A Mack
> University of Rochester
> Cardiovascular Research Institute (CRI)
> 575 Elmwood Avenue, MRBX 2-11001-A
> Rochester, NY 14572
> 585-273-1914
> 
> 
> > ----------
> > From: 	wasielewski.reinhard.von <@t> mh-hannover.de
> > Sent: 	Wednesday, August 24, 2005 8:22 AM
> > To: 	histonet <@t> lists.utsouthwestern.edu
> > Subject: 	[Histonet] VEGF and VEGF-R in FFPE mouse
> > 
> > Dear Histonetters,
> > may I ask your help to find out the best antibody and staining procedure
> > for formalin-
> > fixed, paraffin embedded mouse tissue for the markers VEGF and VEGFR ?
> > 
> > Many thanks in advance - best regards
> > 
> > Reinhard
> > 
> > 
> > 
> > PD Dr. med. Reinhard von Wasielewski
> > 
> > 
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> > 
> > 
> 
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