e-mail+to+Ibii+Takahisa

e-mail to ibii.takahisa@canon.co.jp This e-mail failed to send Hi Takahisa Ibii,

I see that you are the corresponding author for the paper titled "Direct Immobilization of Gold-Binding Antibody". I also see that Hidenori Shiotsuka and Takeshi Imamura are authors of this paper as well. I have some questions for them, but I don't know how to contact them. Could you give me their addressess and/or forward this e-mail to them?

Here are my questions for them. I see that they are the inventors on a patent found here: http://www.freepatentsonline.com/y2008/0206784.html

In this patent document, they use the M13KE vector from New England Biolabs to express an scFv molecule as a fusion with the pIII protein. I also want to express scFv molecules as a fusion with the pIII protein using this M13KE vector, but I wasn't completely sure if this was possible since New England Biolabs only advertises use of peptide-pIII fusions rather than scFv-pIII fusions. They recommend peptides <50 aa (amino acids) whereas the scFv is 250 aa (amino acids). So my questions are: Does it work okay to use scFv with this M13KE vector? If not, is there an alternative commercially available vector you would recommend? Does the infectivity of M13KE phage decrease greatly when a large scFv is attached to the pIII protein? When getting bacteria to express the phage, is it necessary to use a helper phage also?

I also have a question about the restriction sites used. Typically, SfiI and NotI sites have been used to cut scFv genes and ligate them into a phagemid or phage vector. However, the M13KE vector does not have these sites. The M13KE vector uses Acc651-KpnI and EagI sites instead. Did you change the restriction sites of the vector or did you change the restriction sites of your scFv inserts?

Note that my scFv are constructed from the B cells of mice with cancer. This is not necessary for you to know to answer my questions, but I just thought I would let you know anyway (just fyi).

Thanks for any information you can offer me!

Best regards, Kurt Whittemore

Graduate Student Arizona State University BIODESIGN INSTITUTE Center for Innovations in Medicine 1001 S McALLISTER AVE TEMPE, AZ 85287

2nd attempt Corresponding author: kmiz@kuma.che.tohoku.ac.jp

Hi,

I see that you are the corresponding author for the paper titled "Biomimetic Engineering of Modular Bispecific Antibodies for Biomolecule Immobilization". I also see that Hidenori Shiotsuka and Takeshi Imamura are authors of this paper. I have some questions for them, but I don't know how to contact them. Could you give me their e-mail addressess and/or forward this e-mail to them?

Here are my questions for them. I see that they are the inventors on a patent found here: http://www.freepatentsonline.com/y2008/0206784.html

In this patent document, they use the M13KE vector from New England Biolabs to express an scFv molecule as a fusion with the pIII protein. I also want to express scFv molecules as a fusion with the pIII protein using this M13KE vector, but I wasn't completely sure if this was possible since New England Biolabs only advertises use of peptide-pIII fusions rather than scFv-pIII fusions. They recommend peptides <50 aa (amino acids) whereas the scFv is 250 aa (amino acids). So my questions are: Does it work okay to use scFv with this M13KE vector? If not, is there an alternative commercially available vector you would recommend? Does the infectivity of M13KE phage decrease greatly when a large scFv is attached to the pIII protein? When getting bacteria to express the phage, is it necessary to use a helper phage also?

I also have a question about the restriction sites used. Typically, SfiI and NotI sites have been used to cut scFv genes and ligate them into a phagemid or phage vector. However, the M13KE vector does not have these sites. The M13KE vector uses Acc651-KpnI and EagI sites instead. Did you change the restriction sites of the vector or did you change the restriction sites of your scFv inserts?

Note that my scFv are constructed from the B cells of mice with cancer. This is not necessary for you to know to answer my questions, but I just thought I would let you know anyway (just fyi).

Thanks for any information you can offer me!

Best regards, Kurt Whittemore

Graduate Student Arizona State University BIODESIGN INSTITUTE Center for Innovations in Medicine 1001 S McALLISTER AVE TEMPE, AZ 85287