Re: [ccp4bb] Off-topic His-Antibody
Dear All, Thanks for all the replies on and off-board. I received around twenty replies and the majority have spoken in favor of the QIAgen BSA-free anti-5His mAb from QIAGEN. Not to be bias, a couple of people recommended the one from Abcam as well. Thanks again, Dan
Re: [ccp4bb] Off-topic His-Antibody
I have been using an HRP conjugated antibody from Thermo that can detect maybe 3-8ng of HisTagged protein from insect cell lysates. Performance is ok, but with the sensitivity I am not so happy with. How is the sensitivity with the Qiagen antibody? How many picograms you need for a clear band? Does anyone know a good HRP conjugated antibody? Regards, Juha On 26 June 2012 06:59, Luca Jovine luca.jov...@ki.se wrote: We routinely use BSA-free anti-5His mAb from QIAGEN, and it works just fine (at least using media and lysates of both insect and mammalian cells): http://www.qiagen.com/products/protein/detection/qiaexpressdetectionsystem/penta-hisantibody.aspx Good luck, Luca Luca Jovine, Ph.D. Assistant Professor EMBO Young Investigator Karolinska Institutet Department of Biosciences and Nutrition Center for Biosciences Hälsovägen 7, SE-141 83 Huddinge, Sweden Voice: +46.(0)8.524-81136 FAX: +46.(0)8.6081-501 E-mail: luca.jov...@ki.se W3: http://jovinelab.org From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of D Bonsor [ dbon...@ihv.umaryland.edu] Sent: Monday, June 25, 2012 23:33 To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Off-topic His-Antibody Are there any good antibodies for His-tags on the market. I have never used one and I heard several stories that they were poor and not worth the money, over the past few years. The only post I could find on the BB was from 2008. Have His-Antibodies improved or are they still rubbish. -- Genius may have its limitations, but stupidity is not thus handicapped. -Elbert Hubbard *
Re: [ccp4bb] Off-topic His-Antibody
Our experience essentially agrees with QIAGEN's claim that you can detect proteins in the low nanogram range, i.e. 1-2 ng... so not a major improvement over your current antibody, I'm afraid. They also sell an HRP conjugate version: http://www.qiagen.com/products/protein/detection/qiaexpressdetectionsystem/penta-hishrpconjugatekit.aspx but we have never tried it... and now I'll stop here, otherwise you'll start thinking I own QIAGEN shares :-) Luca On Jun 26, 2012, at 09:54 , Juha Vahokoski wrote: I have been using an HRP conjugated antibody from Thermo that can detect maybe 3-8ng of HisTagged protein from insect cell lysates. Performance is ok, but with the sensitivity I am not so happy with. How is the sensitivity with the Qiagen antibody? How many picograms you need for a clear band? Does anyone know a good HRP conjugated antibody? Regards, Juha On 26 June 2012 06:59, Luca Jovine luca.jov...@ki.se wrote: We routinely use BSA-free anti-5His mAb from QIAGEN, and it works just fine (at least using media and lysates of both insect and mammalian cells): http://www.qiagen.com/products/protein/detection/qiaexpressdetectionsystem/penta-hisantibody.aspx Good luck, Luca Luca Jovine, Ph.D. Assistant Professor EMBO Young Investigator Karolinska Institutet Department of Biosciences and Nutrition Center for Biosciences Hälsovägen 7, SE-141 83 Huddinge, Sweden Voice: +46.(0)8.524-81136 FAX: +46.(0)8.6081-501 E-mail: luca.jov...@ki.se W3: http://jovinelab.org From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of D Bonsor [dbon...@ihv.umaryland.edu] Sent: Monday, June 25, 2012 23:33 To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Off-topic His-Antibody Are there any good antibodies for His-tags on the market. I have never used one and I heard several stories that they were poor and not worth the money, over the past few years. The only post I could find on the BB was from 2008. Have His-Antibodies improved or are they still rubbish. -- Genius may have its limitations, but stupidity is not thus handicapped. -Elbert Hubbard *
Re: [ccp4bb] Off-topic His-Antibody
I'll just second, third, and fourth the assessment of the Qiagen anti-pentaHis antibody. I pretty much gave up on anti-His antibodies until we tried this out in the lab and now everyone loves it. Reasonable sensitivity and specificity (we use IR dye-conjugated secondaries and a Li-Cor Odyssey scanner for signal detection) and not bad on cost. And it does detect hexa- and decahistidine tags. Not using HRP anymore since we switched over to IR detection a few years ago. HTH- Brad On Tue, Jun 26, 2012 at 4:17 AM, Luca Jovine luca.jov...@ki.se wrote: Our experience essentially agrees with QIAGEN's claim that you can detect proteins in the low nanogram range, i.e. 1-2 ng... so not a major improvement over your current antibody, I'm afraid. They also sell an HRP conjugate version: http://www.qiagen.com/products/protein/detection/qiaexpressdetectionsystem/penta-hishrpconjugatekit.aspx but we have never tried it... and now I'll stop here, otherwise you'll start thinking I own QIAGEN shares :-) Luca On Jun 26, 2012, at 09:54 , Juha Vahokoski wrote: I have been using an HRP conjugated antibody from Thermo that can detect maybe 3-8ng of HisTagged protein from insect cell lysates. Performance is ok, but with the sensitivity I am not so happy with. How is the sensitivity with the Qiagen antibody? How many picograms you need for a clear band? Does anyone know a good HRP conjugated antibody? Regards, Juha On 26 June 2012 06:59, Luca Jovine luca.jov...@ki.se wrote: We routinely use BSA-free anti-5His mAb from QIAGEN, and it works just fine (at least using media and lysates of both insect and mammalian cells): http://www.qiagen.com/products/protein/detection/qiaexpressdetectionsystem/penta-hisantibody.aspx Good luck, Luca Luca Jovine, Ph.D. Assistant Professor EMBO Young Investigator Karolinska Institutet Department of Biosciences and Nutrition Center for Biosciences Hälsovägen 7, SE-141 83 Huddinge, Sweden Voice: +46.(0)8.524-81136 FAX: +46.(0)8.6081-501 E-mail: luca.jov...@ki.se W3: http://jovinelab.org From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of D Bonsor [ dbon...@ihv.umaryland.edu] Sent: Monday, June 25, 2012 23:33 To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Off-topic His-Antibody Are there any good antibodies for His-tags on the market. I have never used one and I heard several stories that they were poor and not worth the money, over the past few years. The only post I could find on the BB was from 2008. Have His-Antibodies improved or are they still rubbish. -- Genius may have its limitations, but stupidity is not thus handicapped. -Elbert Hubbard *
Re: [ccp4bb] Off-topic His-Antibody
The one we have is rubbish. I can tell a vendor name if you want off-list. A lot of non-specific bands + high background level makes it almost impossible to use against cell lysate. It sort of works against purified His-tag protein, but not perfect either. On Mon, Jun 25, 2012 at 4:33 PM, D Bonsor dbon...@ihv.umaryland.edu wrote: Are there any good antibodies for His-tags on the market. I have never used one and I heard several stories that they were poor and not worth the money, over the past few years. The only post I could find on the BB was from 2008. Have His-Antibodies improved or are they still rubbish.
Re: [ccp4bb] Off-topic His-Antibody
We routinely use BSA-free anti-5His mAb from QIAGEN, and it works just fine (at least using media and lysates of both insect and mammalian cells): http://www.qiagen.com/products/protein/detection/qiaexpressdetectionsystem/penta-hisantibody.aspx Good luck, Luca Luca Jovine, Ph.D. Assistant Professor EMBO Young Investigator Karolinska Institutet Department of Biosciences and Nutrition Center for Biosciences Hälsovägen 7, SE-141 83 Huddinge, Sweden Voice: +46.(0)8.524-81136 FAX: +46.(0)8.6081-501 E-mail: luca.jov...@ki.se W3: http://jovinelab.org From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of D Bonsor [dbon...@ihv.umaryland.edu] Sent: Monday, June 25, 2012 23:33 To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Off-topic His-Antibody Are there any good antibodies for His-tags on the market. I have never used one and I heard several stories that they were poor and not worth the money, over the past few years. The only post I could find on the BB was from 2008. Have His-Antibodies improved or are they still rubbish.
Re: [ccp4bb] Off-topic His-Antibody
We also use that same antibody, and are satisfied overall. On 6/25/12 8:59 PM, Luca Jovine wrote: We routinely use BSA-free anti-5His mAb from QIAGEN, and it works just fine (at least using media and lysates of both insect and mammalian cells): http://www.qiagen.com/products/protein/detection/qiaexpressdetectionsystem/penta-hisantibody.aspx Good luck, Luca Luca Jovine, Ph.D. Assistant Professor EMBO Young Investigator Karolinska Institutet Department of Biosciences and Nutrition Center for Biosciences Hälsovägen 7, SE-141 83 Huddinge, Sweden Voice: +46.(0)8.524-81136 FAX: +46.(0)8.6081-501 E-mail: luca.jov...@ki.se W3: http://jovinelab.org From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of D Bonsor [dbon...@ihv.umaryland.edu] Sent: Monday, June 25, 2012 23:33 To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Off-topic His-Antibody Are there any good antibodies for His-tags on the market. I have never used one and I heard several stories that they were poor and not worth the money, over the past few years. The only post I could find on the BB was from 2008. Have His-Antibodies improved or are they still rubbish. -- Engin Özkan Post-doctoral Scholar Laboratory of K. Christopher Garcia Howard Hughes Medical Institute Dept of Molecular and Cellular Physiology 279 Campus Drive, Beckman Center B173 Stanford School of Medicine Stanford, CA 94305 w ph: (650)-498-7111 cell: (650)-862-8563
