IGEM:MIT/2005/Antibodies

http://www.ci.berkeley.ca.us/environmentalhealth/images/bacteria.gif

TO DO LIST
1) Obtain plasmid with anti-rabbit scFV from authors of the SCFV antibody Piezoimmunosensors paper
 * Zeng@oakland.edu
 * Ask
 * NCBI sequence different from sequence in paper
 * conserved region
 * get clone on plasmid

2) Determine and order fluorescein - tagged DNA for use in determining dimerization distance

3) Blast and check the "low complexity" region of the light chain in the scFV
 * confirm the match to rice firus is a fluke!

4) Check 3D structure of antibodies (ask Natalie and Kate)

5) Additional info: swiss-prot

Goals

 * Find an antigen and its antibodies
 * Sequence of the fusing antibody
 * Proper distance between epitopes

Resources

 * Post-translational Modification
 * Antibody Resources
 * cDNA banks/libraries

Possible Sequences for Secondary Antibody
 Anti-rabbit 
 * Sequence: Sequence of synthetic construct anti-rabbit IgG scFV mRNA, partial cds
 * Paper: scFv Antibody Piezoimmunosensors
 * Summary: ../Summary scFvSensor/

 Anti-mouse 
 *  ScFVs: single chain fragment variable (we shouldn't have to engineer these too much) 
 * Sequence anti-mouse1: Synthetic construct clone H2 anti-mouse L1 scFv antibody mRNA, partial cds.
 * Sequence anti-mouse2: Synthetic construct clone G6 anti-mouse L1 scFv antibody mRNA, partial cds.
 * Paper covering anti-mouse1 and 2: Single chain Fv antibodies against neural cell adhesion molecule L1 trigger L1 functions in cultured neurons
 * Summary of anti-mouse1 and 2 paper: ../Summary anti-mouse 1 and 2/


 * Sequence of anti-mouse3: Synthetic construct clone C12 anti-mouse CHL1 scFv antibody mRNA, partial cds.
 * Sequence of amti-mouse4: Synthetic construct clone 6C2 anti-mouse CHL1 scFv antibody mRNA, partial cds.
 * Paper covering anti-mouse3 and 4: Single chain variable fragment antibodies against the neural adhesion molecule CHL1 (close homolog of L1) enhance neurite outgrowth
 * Summary of anti-mouse3 and 4: ../Summary anti-mouse 3 and 4/


 *  Light and Heavy Chains (we will need to engineer a linker sequence for these sequences) 
 * Rattus norvegicus anti-mouse IL-4 specific monoclonal antibody 11B11 light chain variable region mRNA, partial cds
 * Rattus norvegicus anti-mouse IL-4 specific monoclonal antibody 11B11 heavy chain variable region mRNA, partial cds.
 * Mus musculus anti-mouse IgDa immunoglobulin light chain variable region mRNA, partial cds.
 * Mus musculus anti-mouse IgDa immunoglobulin heavy chain variable region mRNA, partial cds.

Papers

 * Protein engineering of antibody binding sites (1988)
 * ../Summary scFv/

Books

 * Sequences of Proteins of Immunological Interest, Kabat. Call Number: QD431.S59 1991.
 * [[Image:Kabat91.pdf]]


 * Kabat Database


 * Kabat Database Site -- requires payment


 * Kabat Database Searcher -- free


 * Antibody Engineering: A practical approach, McCafferty et al. 1996


 * [[Image:Filpula96.pdf|Filpula's discussion of the design of the scFV proteins; includes a discussion of linker designs and references to places where the various ones are being used.]]


 * [[Image:Johnson96.pdf|Johnson's discussion describing ways to determine the Kd (dissociation constant) of the manufactured antibodies.]]


 * [[Image:Pluckthun96.pdf|Pluckthun's discussion of how to engineer excretion of antibodies and production of large amounts in E. coli.]]


 * [[Image:Pope96.pdf|Pope's discussion detailing how the anti-rabbit scFv protein Maxine discovered yesterday was designed and made.]]

Questions, Comments, Concerns, Suggestions

 * R.M. de Wildt, C.R. Mundy, B.D. Gorick and I.M. Tomlinson, Antibody arrays for high-throughput screening of antibody–antigen interactions. Nat. Biotechnol. 18 (2000), pp. 989–994.
 * ../Http://www.sciencedirect.com/science? ob=ArticleURL& udi=B6T1G-44BFHS3-4Y8& user=501045& coverDate=01%2F01%2F1900& fmt=summary& orig=search& qd=1& cdi=4890&view=c& acct=C000022659& version=1& urlVersion=0& userid=501045&md5=31926e4c3e8e19fbea8076163f7dc6a3&ref=full Abstract-EMBASE/
 * ../Http://www.sciencedirect.com/science? ob=ArticleURL& udi=B6WR5-44C90DM-2W6& user=501045& coverDate=01%2F01%2F1900& fmt=summary& orig=search& qd=1& cdi=7009&view=c& acct=C000022659& version=1& urlVersion=0& userid=501045&md5=bb68434e190bccde0798e4939684f6b7&ref=full Abtract-Elsevier BIOBASE/
 * ../Http://www.sciencedirect.com/science? ob=ArticleURL& udi=B6WVB-45DHTFR-4FV& user=501045& coverDate=01%2F01%2F1900& fmt=summary& orig=search& qd=1& cdi=7098&view=c& acct=C000022659& version=1& urlVersion=0& userid=501045&md5=aecd12dab51d29e884ca65f7c41edc99&ref=full Abstract-MEDLINE/
 * ../Http://www.sciencedirect.com/science? ob=RedirectURL& method=outwardLink& partnerName=3& targetURL=http%3A%2F%2Fdx.doi.org%2F10.1038%2F79494& acct=C000022659& version=1& userid=501045&md5=0b0cdc494a012ba14ea1e5d9f685d651 Full Text-CrossRef/

Companies checked for antibodies:
eBioscience: no

Anit-Florouscine

 * sequence Substantial Energetic Improvement with Minimal Strctural Perturbation in a High Affinity Mutant Antibody
 * long paper. used as a reference for scFv of antiflourescine types 4-4-20 and 4m5.3