American Society of Hematology

Fig. 9.

$Fig. 9. Detection of WNT5a protein. / (A) Expression of WNT5a protein in stably transfected QCE6 cells. Both lanes contain 25 μg of protein from culture lysates of either stably transfected WNT5a/QCE6 (lane 1) or parental QCE6 cells (lane 2), which were subsequently blotted with antibody reactive to themyc-fusion tag. Arrow indicates the position of an approximately 50-kd band that is present only in the stably transfected cells, which corresponds to the WNT5a fusion protein. (B) Partial purification of WNT5a protein from WNT5a-conditioned media. This shows equal volumes of either nonpurified WNT5a CM (lane 3), the effluent fraction of WNT5a CM run over an Ni-NTA column (lane 4), or the nickel-binding fraction eluted from the same column (lane 5). An arrow indicates the position of an approximately 50-kd WNT5a band that specifically bound to the Ni-NTA column owing to the multiple histidine residues of the fusion protein tag. The protein concentrations of WNT5a CM and Ni-NTA column–purified WNT5a–containing fraction were 14 μg/μL and 52 ng/μL, respectively.$

Detection of WNT5a protein.

(A) Expression of WNT5a protein in stably transfected QCE6 cells. Both lanes contain 25 μg of protein from culture lysates of either stably transfected WNT5a/QCE6 (lane 1) or parental QCE6 cells (lane 2), which were subsequently blotted with antibody reactive to themyc-fusion tag. Arrow indicates the position of an approximately 50-kd band that is present only in the stably transfected cells, which corresponds to the WNT5a fusion protein. (B) Partial purification of WNT5a protein from WNT5a-conditioned media. This shows equal volumes of either nonpurified WNT5a CM (lane 3), the effluent fraction of WNT5a CM run over an Ni-NTA column (lane 4), or the nickel-binding fraction eluted from the same column (lane 5). An arrow indicates the position of an approximately 50-kd WNT5a band that specifically bound to the Ni-NTA column owing to the multiple histidine residues of the fusion protein tag. The protein concentrations of WNT5a CM and Ni-NTA column–purified WNT5a–containing fraction were 14 μg/μL and 52 ng/μL, respectively.

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