How to choose the correct antibody？
1，How to choose the correct antibody？
Target of interest
• learn as much as you can about the target protein you are testing for.
• know the full name of the protein you are plan to detect
• search the target protein on Uniprot (www.uniport.org) and make sure the ID of the protein.
2. Uniport
For example：
http://www.uniprot.org/uniprot/P00533
3. Protein Target
• If your target protein has many isoforms, you should make sure the antibody could recognized your isoform. Some antibody supplier has the immune sequence or N-term, C-term informations on their datasheet.
Know your samples as much as possible!
Do you know the expressed level of the target protein? Do you have RNA data or mass spectrometric data for your samples?
• You can search the RNA data here: www.proteinatlas.org
4. Species/Reactivity
• When you search the antibody for your target protein, you could see the species that list in the manual such as: H, M, R, B, D, Mk, P, S, Z and so on.
• If you couldn’t get the species you need, please contact with our staff for more information, we could compare the sequence and the homology of the corresponding species, and then determine whether it can be detect.
5. Application
Common application including: ELISA, WB, IHC, IF, ICC, IP, FC, ChIP
• If the application didn’t list on the manual, it don’t mean this kit can’t do this experiment, sometimes maybe we didn’t detect in our labor other reasons, if you want more application informations please contact our staff.
6. Polyconal antibody or monoclonal antibody?
• Due to batch to batch consistency, each monoclonal antibody recognizes a single epitope, whose specificity is determined by mAb。
• polyclonal antibody recognizes more epitope and provide more application compatibility.
7. The advantage and disadvantage for polyclonal antibody
Advantage:
• Help amplify the signal of target protein with low express signal.
• Can recognize more epitope, so it can get better result in IP/ChIP.
• It is more tolerant of small changes of antigens than monoclonal antibody.
• It can recognize the protein which has high homology to immunogenic protein
• Polyclonal antibodies are usually the first choice for detecting denatured proteins
• Multiple epitopes， it often provide more robust detection
Disadvantage:
•It’s easy to produce batch difference.
•It can easy Induce non-specific antibodies, that may produce background signals in some applications.
• You should confirm with us, whether there’s any cross-reactive.
8. The advantage and disadvantage for monoclonal antibody
Advantage:
•All of the batches are the same, that would ensure the consistency experiment procedure and results.
• Monoclonal antibodies produce low background staining
• The reproducibility of monoclonal antibody results is very high
Disadvantage:
• Too specific
• It can only bound one sit, however the processed antigen always missing some epitope so it’s won’t reactive.
• The quantity is small and the production cost is high
9，The host we often use for production antibodies
• Mouse Rat（monoclonal antibody）
• Rabbit（polyclonal antibody，Monoclonal antibody）
• Goat Sheep（polyclonal antibody）
10，The immunogen we used for production antibody
polypeptide
Antibodies with high specificity and low cross reaction were prepared by the design of epitopes：
The antibody can be used for: ELISA， WB，IHC，ICC and so on.
Antibodies that modify proteins such as phosphorylation, acetylation, methylation.
Recombinant protein
Preparation of antibodies against different epitopes: Recombinant proteins are usually expressed in E. coli, and mammalian proteins still differ in glycosylation and folding.
11, Secondary antibody
type：IgA IgG IgM IgY
label：Biotin, HRP, AP, AMCA, Cy3, FITC, TRITC
Alexa Fluor 350, 405, 488, 532, 555, 568, 594, 633, 647, 660, 680, 750, 790
Target of interest
• learn as much as you can about the target protein you are testing for.
• know the full name of the protein you are plan to detect
• search the target protein on Uniprot (www.uniport.org) and make sure the ID of the protein.
2. Uniport
For example：
http://www.uniprot.org/uniprot/P00533
3. Protein Target
• If your target protein has many isoforms, you should make sure the antibody could recognized your isoform. Some antibody supplier has the immune sequence or N-term, C-term informations on their datasheet.
Know your samples as much as possible!
Do you know the expressed level of the target protein? Do you have RNA data or mass spectrometric data for your samples?
• You can search the RNA data here: www.proteinatlas.org
4. Species/Reactivity
• When you search the antibody for your target protein, you could see the species that list in the manual such as: H, M, R, B, D, Mk, P, S, Z and so on.
• If you couldn’t get the species you need, please contact with our staff for more information, we could compare the sequence and the homology of the corresponding species, and then determine whether it can be detect.
5. Application
Common application including: ELISA, WB, IHC, IF, ICC, IP, FC, ChIP
• If the application didn’t list on the manual, it don’t mean this kit can’t do this experiment, sometimes maybe we didn’t detect in our labor other reasons, if you want more application informations please contact our staff.
6. Polyconal antibody or monoclonal antibody?
• Due to batch to batch consistency, each monoclonal antibody recognizes a single epitope, whose specificity is determined by mAb。
• polyclonal antibody recognizes more epitope and provide more application compatibility.
7. The advantage and disadvantage for polyclonal antibody
Advantage:
• Help amplify the signal of target protein with low express signal.
• Can recognize more epitope, so it can get better result in IP/ChIP.
• It is more tolerant of small changes of antigens than monoclonal antibody.
• It can recognize the protein which has high homology to immunogenic protein
• Polyclonal antibodies are usually the first choice for detecting denatured proteins
• Multiple epitopes， it often provide more robust detection
Disadvantage:
•It’s easy to produce batch difference.
•It can easy Induce non-specific antibodies, that may produce background signals in some applications.
• You should confirm with us, whether there’s any cross-reactive.
8. The advantage and disadvantage for monoclonal antibody
Advantage:
•All of the batches are the same, that would ensure the consistency experiment procedure and results.
• Monoclonal antibodies produce low background staining
• The reproducibility of monoclonal antibody results is very high
Disadvantage:
• Too specific
• It can only bound one sit, however the processed antigen always missing some epitope so it’s won’t reactive.
• The quantity is small and the production cost is high
9，The host we often use for production antibodies
• Mouse Rat（monoclonal antibody）
• Rabbit（polyclonal antibody，Monoclonal antibody）
• Goat Sheep（polyclonal antibody）
10，The immunogen we used for production antibody
polypeptide
Antibodies with high specificity and low cross reaction were prepared by the design of epitopes：
The antibody can be used for: ELISA， WB，IHC，ICC and so on.
Antibodies that modify proteins such as phosphorylation, acetylation, methylation.
Recombinant protein
Preparation of antibodies against different epitopes: Recombinant proteins are usually expressed in E. coli, and mammalian proteins still differ in glycosylation and folding.
11, Secondary antibody
type：IgA IgG IgM IgY
label：Biotin, HRP, AP, AMCA, Cy3, FITC, TRITC
Alexa Fluor 350, 405, 488, 532, 555, 568, 594, 633, 647, 660, 680, 750, 790
