Author Topic: Please help me to understand!  (Read 2055 times)

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Offline drliem

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Please help me to understand!
« on: February 20, 2011, 10:19:23 PM »
Dear every body,
I am PhD student the first year.
This is the first time I study IHC.
I read some documents. I do not understand about
- How is a positive control slide? What for?
- How is a negative control slide? What for?
- How is a reagent control slide? What for?

Please held me to understand.
I am looking forward to hearing you.
My email is bsliem74@gmail.com

Best regards

Pham Hieu LIEM

Please help me to understand!
« on: February 20, 2011, 10:19:23 PM »

Offline excalibur

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Re: Please help me to understand!
« Reply #1 on: February 21, 2011, 08:43:01 AM »
Paula K. Pierce, HTL(ASCP)HT
Excalibur Pathology, Inc.
8901 S Santa Fe, Suite G
Oklahoma City, OK 73139
405-759-3959
www.excaliburpathology.com

Offline pandanac

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Re: Please help me to understand!
« Reply #2 on: March 12, 2011, 08:16:26 AM »
Hi Guys:

 I would like to explain mine..I am looking to map the C-fos in the V1 of the rat brain.

1. I use Brain tissue slices, PFA perfused, Fixed, sectioned and mounted on slides and frozen until further use.

2. Used TBS-twn-20 for washing steps.

3. Nonspecific blocking for 2hrs by serum (10%).

4. Primary antibody I used first was from Santa Cruz Dilution 1:100 overnight incubation

5. Secondary antibody Alexa Fluor-594 dilution 1:300 incubation for 2 hrs at room temperature

6. Washed several times and add the DAPI incubated for 20 min

7. Washed, added the antifade and cover slipped.

the problem is I am seeing positive neurons (mean the antibody is binding to neurons which can be seen in the control tissue, which is from the rat not exposed to any visual stimulation [that's what i do to induce the c-fos in the V1 area f the brain ])

I changed the antibody from santa cruz to ABcam antibody, however I am still seeing the same result :-(

can any one help me in solving this problem.

Offline REssy

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Re: Please help me to understand!
« Reply #3 on: July 13, 2011, 02:11:26 AM »
excalibur shared literally marvelous link. Thanks a lot.

Re: Please help me to understand!
« Reply #3 on: July 13, 2011, 02:11:26 AM »