Hello Gail,

Your message came in just as I was about to send the same request. I am 
having the same problem and nothing I have tried has worked either. I am 
at my wits end.
I am using several different antibodies and 4 different alexa fluors, 
rabbit and mouse, all on rat brain sections. I ran a no primary control, 
a no secondary control and a section with both and they all three looked 
the same! And it was punctate, it looked so real!
I took a section that had only been it PBS and saw nothing.
If what you say is true about the blocking serum, you have made my day.
We never thought of that.
I will have to start over and redo three weeks of work, but at least it 
will be accurate.
Hopefully someone out there can help us. I think most of them are at NSH 
so you may need to send this out again.

Kathleen
On Tuesday, October 1, 2002, at 11:39 AM, Gail Donegan wrote:

> Hello All,
>
> I am staining for human extracellular matrix proteins using mouse
> anti-collagen/fibronectin/elastin etc. I am using a 
> Fluorscein-conjugated
> goat anti-mouse secondary antibody. Despite various preventative 
> methods I
> am getting perfect staining on my negative (or null) control which has 
> no
> primary added.
>
> I've used various dilutions, Bovine, human and rabbit serum block, and
> multiple blocking steps
>
> all with no improvement. If anything, the secondary has greater affinity
> for matrix proteins than it does for the IgG part of the primary. I'm
> wondering whether, since the serum itself has matrix proteins in it,
> whether to completely forego the blocking step.
>
>
> Any thoughts or ideas extremely welcome,
>
> Thanks
>
> Gail
>
>
>
> Gail Donegan
> Department of Clinical Engineering,
> The University of Liverpool,
> Duncan Building,
> Daulby Street,
> Liverpool,
> L69 3GA.
> U.K.
>
> Tel: +44(0)151 706 5198
> Fax: +44(0)151 706 5803
>
> .
>
> Gail Donegan
> Department of Clinical Engineering,
> The University of Liverpool,
> Duncan Building,
> Daulby Street,
> Liverpool,
> L69 3GA.
> U.K.
>
> Tel:    +44(0)151 706 5198
> Fax:    +44(0)151 706 5803
>
>