| Polyclonal antisera |
| | Q: | Are polyclonal antibodies or monoclonal antibodies more suitable? |
| A: | Please see the following overview:
| | Advantages | Disadvantages | | Polyclonal antibodies | inexpensive | limited availability | | | short preparation times (normally 6-8 weeks) | mixture of antibodies having differing properties (constants, epitope binding) | | | | cannot be reproduced or can only be reproduced with difficulty | | Monoclonal antibodies | always reproducible and can be prepared in any desired quantity | cost-intensive preparation | | | possibility of selecting antibodies which possess very specific properties | processing time of up to 6 months |
|
| | Q: | Which animals get immunised? |
| A: | - rabbit
- goat
- mouse
- guinea pig
- rat
other species on request |
| | Q: | What kind of antigens can be used? |
| A: |
- protein
- peptide
- peptide conjugate
- hapten
- small molecule
|
| | Q: | In what form the antigen can be supplied for immunisation? |
| A: |
- hyophilised
- dissolved or precipitated in buffer (concentration 0.5 - 1mg/ml)
- in polyacrylamide gel - Coomassie-staining is possible; as pieces which are pH-neutral (as a result of being washed 2-3x in water) and kept moist (in distilled water), up to a maximum of 500 µl of swollen gel per injection
- nitrocellulose
|
| | Q: | What should be the purity of the antigens? |
| A: | - proteins: >95%
- peptides: 70-80%
|
| | Q: | What quantities of antigen are required per animal? |
| A: |
Immunisation in
|
Goat
|
Rabbit
|
|
Mouse
|
| |
|
|
|
|
|
Peptide
|
5 mg
|
2 mg
|
|
1 mg
|
|
|
|
|
|
|
|
Protein solution
|
2,5 mg
|
0,5-1,5 mg
|
|
0,5 mg
|
|
|
|
|
|
|
|
Protein in gel
|
1 mg
|
0,7 mg
|
|
-
|
|
|
|
|
|
|
|
Peptide conjugate
|
7 mg
|
5 mg
|
|
2 mg
|
|
|
|
|
|
|
|
Nitrocellulose
|
|
|
|
|
The prospects of obtaining a good immune response increase in proportion to the amount of antigen present. For affinity purification we need 3- 6 mg of antigen. If other quantities or concentrations are available, please contact us. We shall find a solution. |
| | Q: | What information with regard to the antigen do we have to have? |
| A: | Protein:- buffer
- concentration
- purity
- molecular weight
Native protein: source
Recombinant protein: fusion moiety and expression system
Peptide:
Peptide conjugate:- quantity
- purity
- sequence
- protein carrier
Hapten:
Oligo- or polysaccarides:- Structure
- Source
- Purity
- Quantity
|
| | Q: | Which buffer contents or antigen contaminations are acceptable for immunisation? |
| A: | The following additives in buffers can be accepted if the concentration of the antigen is ca.1mg/ml: (maximum)
- 1M Urea
- 200 mM Imidazole (in this case without NaCl)
- 1 mM EDTA
- 2 mM DTT (Dithiotriethole)
- 300 mM NaCl (in this case without Imidazole)
- 0,1% Maltose, Manitol and Trehalose
- 0,1% SDS
- 10% Glycerol
|
| | Q: | What is the chronological sequence of an immunisation? |
| A: | There are standard protocols for the immunisation (see the pages concerning polyclonal antibodies). It is possible to vary them in accordance with the customer’s wishes, and they depend on the quantity and nature of the antigen. |
| | Q: | How long can the animals be kept? |
| A: | In most cases it is possible to keep animals as long as the customer needs. The customer decides how long the animals are to be kept and when they are to be final bleeded or disposed of. On every occasion the antisera are supplied the customer is given a fax reply for communicating the further requirements with regard to animal maintenance. |
printable version