Go Back   Science Forums Biology Forum Molecular Biology Forum Physics Chemistry Forum > Molecular Research Topics Forum > Protein Science
Register Search Today's Posts Mark Forums Read

Protein Science Post Questions and Discuss Protein Structure and Function, Protein Localization etc.. Please do not post Proteomic questions here, there is a Proteomic Forum below.


IGF_1 Detection

IGF_1 Detection - Protein Science

IGF_1 Detection - Post Questions and Discuss Protein Structure and Function, Protein Localization etc.. Please do not post Proteomic questions here, there is a Proteomic Forum below.


Reply
 
LinkBack Thread Tools Display Modes
  #1  
Old 12-24-2006, 01:06 AM
Pipette Filler
Points: 1,655, Level: 24 Points: 1,655, Level: 24 Points: 1,655, Level: 24
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Dec 2006
Posts: 18
Thanks: 0
Thanked 0 Times in 0 Posts
Default IGF_1 Detection



I tried western for IGF-1 twice, but dint find any bands. THe MW is 17.

I am unsure if there is no IGF_1 in the sample or if I should try to optmize my westerns. Is it possible that if the levels can become too low of a protein that yu might not be able to see it even in a control group? Or you should see band, atleast a light one, for wahetevr protein you look for in your tissue?

Thanks
Reply With Quote
  #2  
Old 12-27-2006, 04:44 PM
moleculardude's Avatar
M.D/Ph.D
 
Join Date: Jul 2006
Location: Melbourne Australia
Posts: 423
Thanks: 5
Thanked 10 Times in 8 Posts
Default Re: IGF_1 Detection

Dear Anoopbal,

Have you tried Immunoprecipitation? That way you could pull down all the IGF-1 and then do western blot on a more concentrated sample.

Also make sure you lyse in the smallest volume possible and load most of it.

Before you do all that however, I would make sure you can see IGF-1 with a control using your antibody.

cheers!
Reply With Quote
  #3  
Old 08-29-2013, 11:01 AM
Zagami's Avatar
Ph.D Doctorate
Points: 4,317, Level: 44 Points: 4,317, Level: 44 Points: 4,317, Level: 44
Activity: 33% Activity: 33% Activity: 33%
 
Join Date: Sep 2009
Location: Tabaka Mission Hospital - Kiisi -Kenya & Italy Sicily
Posts: 167
Thanks: 2
Thanked 28 Times in 24 Posts
Default Re: IGF_1 Detection

Insulin-like growth factors (IGFs) belong to a family of peptides that share a high degree of structural homology with insulin. IGFs (IGF-I and IGF-II) have important mitogenic and anabolic actions that are mediated by their binding to specific high affinity cell surface receptors. IGF-I and IGF-II are produced in multiple tissues and are present in blood and other biological fluids in association with specific high affinity IGF binding proteins (IGFBPs). For determination of IGFs it's required their dissociation from IGFBPs. Procedures for direct measurement of IGFs by sample acidification to a pH of about 2.0 to dissociate the complexes followed by neutralization to slightly alkaline pH prior to ELISA or Western blot analysis. The extraction protocol is realized using 20 Ál of EDTA-K3 whole blood spotted on filter paper disc, obtained employing two-hole (filebinder) hole punch with 5.5-6.5 diameter and dried at room temperature for 16 hrs, that must be put in polystyrene tube in which 200 Ál of extraction solution (Tris-HCl buffer 0.005 M at pH 7.0, containing 0.5 ml/L Tween-20) is added and incubate shaking at room temperature for 1 h to allow complete elution of IGF from the filter paper. To the extract is then added 200 Ál of acidification solution (glycine-HCl buffer 0.4 M at pH 2.0) and incubated as above for 30 min, and neutralized by adding 400 Ál of neutralization solution (0.1% sodium dodecylsulfate in 0.85 M of Tris). The extract is mixed for 2 min and used for IGF-I or IGF-II analysis.
Reply With Quote
Reply

Tags
detection , igf1


Thread Tools
Display Modes

Posting Rules
You may not post new threads
You may not post replies
You may not post attachments
You may not edit your posts

BB code is On
Smilies are On
[IMG] code is On
HTML code is Off
Trackbacks are On
Pingbacks are On
Refbacks are On

Forum Jump

Similar Threads
Thread Thread Starter Forum Replies Last Post
Determination of oxytocin in biological samples by isocratichigh-performance liquid chromatography with coulometric detection using C18solid-phase extraction and polyclonal antibody-based immunoaffinity columnpurification tocopherol Forum Chemie 1 10-10-2012 02:11 PM
Monomeric Abeta detection in brain of APP tg mice using westernblotting Stanley Cheung Protocols and Methods Forum 0 05-15-2009 11:17 AM
Determination of oxytocin in biological samples by isocratichigh-performance liquid chromatography with coulometric detection using C18solid-phase extraction and polyclonal antibody-based immunoaffinity columnpurification tocopherol Protocols and Methods Forum 0 11-29-2008 04:24 PM
Determination of oxytocin in biological samples by isocratichigh-performance liquid chromatography with coulometric detection using C18solid-phase extraction and polyclonal antibody-based immunoaffinity columnpurification tocopherol Cell Biology and Cell Culture 0 11-29-2008 04:00 PM
Determination of oxytocin in biological samples by isocratichigh-performance liquid chromatography with coulometric detection using C18solid-phase extraction and polyclonal antibody-based immunoaffinity columnpurification tocopherol Protein Forum 0 03-21-2008 07:38 PM


All times are GMT. The time now is 07:21 AM.


Powered by vBulletin® Version 3.8.4
Copyright ©2000 - 2014, Jelsoft Enterprises Ltd.
Copyright 2005 - 2012 Molecular Station | All Rights Reserved
Page generated in 0.15820 seconds with 16 queries