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More than 10 min exposure time in chemiluminescence detection

chemiuminescence exposure time

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#1 Meg P. Anula

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Posted 05 February 2016 - 07:35 AM

Hi,

 

1. Have anyone experienced using auto-exposure setting in Biorad's ChemiDoc XRS and found that it took longer than 10 min but still no results shown? 

 

Does that mean there's nothing on the membrane?

 

 

2. I plan to reprobe the same membrane with lower dilution of primary antibody. But I am afraid since the membrane has been dried for more than half an hour in the ChemiDoc's darkroom hood while I was testing the exposure time. Would there any effect on the protein that makes it unable to be reprobed?

 

Thanks in advance. 


Edited by Meg P. Anula, 05 February 2016 - 07:37 AM.


#2 bob1

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Posted 07 February 2016 - 01:08 AM

I have worked with an XRS before - basically as far as I can tell, the XRS is a terrible system with very low sensitivity. I have had signal that is easily seen on a 10 min exposure of film (and odyssesy systems) that was not seen even with a 30 min exposure on an XRS. So basically there may be signal there, just the system isn't picking it up.

 

2) The protein will probably be fine, in fact drying a membrane can improve signal in many cases. If you wish to prevent drying, you can sandwich the membrane in cling-wrap or between two plastic sheets.



#3 Meg P. Anula

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Posted 09 February 2016 - 12:59 AM

I have worked with an XRS before - basically as far as I can tell, the XRS is a terrible system with very low sensitivity. I have had signal that is easily seen on a 10 min exposure of film (and odyssesy systems) that was not seen even with a 30 min exposure on an XRS. So basically there may be signal there, just the system isn't picking it up.

 

2) The protein will probably be fine, in fact drying a membrane can improve signal in many cases. If you wish to prevent drying, you can sandwich the membrane in cling-wrap or between two plastic sheets.

 

There're these few bands that I found at the bottom of the membrane upon 30 second exposure. I know it's not the target protein since the negative control also has the band. 

 

Do you know what could it be? 

 

Attached is the picture.chemi 30s.jpg






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