en Bionanotechnology Bionanotechnology مقاله پژوهشی Original Research Article <span style="font-size:10pt"><span new="" roman="" style="font-family:" times=""><b><span style="font-size:9.0pt"><span style="background:#2d7f8f"><span style="color:white">Background & Objective</span></span></span></b><span style="font-size:9.0pt"><span style="background:#2d7f8f"><span style="color:white">:</span></span></span> &nbsp;<span style="font-size:9.0pt"><span style="color:black">Breast cancer is an international concern due to its high prevalence worldwide. Human epidermal growth<b> </b>factor receptor 2 (HER2)-positive breast cancers were observed in 15-20% of breast cancers, with higher aggressiveness and poor prognosis. The single-chain fragment variable (scFv) structure includes variable regions of antibody light and heavy chains. Despite the promising potential of anti-HER2 scFv for non-invasive detection of HER2 status, its low expression and solubility remain significant challenges. Here, we intended to optimize the<b> </b>anti-HER2 scFv expression and solubility in BL21-pET-22 (anti-HER2 scFv).<b> </b></span></span></span></span><br> <span style="font-size:10pt"><span new="" roman="" style="font-family:" times=""><b><span style="font-size:9.0pt"><span style="background:#2d7f8f"><span style="color:white"><span style="letter-spacing:-.1pt">&nbsp;Materials & Methods:</span></span></span></span></b> &nbsp;<span style="font-size:9.0pt"><span style="color:black"><span style="letter-spacing:-.1pt">The effects of ethanol stress, aeration, post-induction temperatures, induction-starting times, culture mediums, and MgCl₂ addition were examined.</span></span></span></span></span><br> <span style="font-size:10pt"><span new="" roman="" style="font-family:" times=""><b><span lang="EN-IN" style="font-size:9.0pt"><span style="background:#2d7f8f"><span style="color:white">Results: </span></span></span></b><b>&nbsp;</b><span style="font-size:9.0pt"><span style="color:black"><span style="letter-spacing:-.1pt">We observed that ethanol stress (1% v/v) increased protein expression and decreased protein solubility at 37 &deg;C. However, it enhanced anti-HER2 scFv solubility at lower post-induction temperature (22 &deg;C). Induction at OD₆₀₀=1 elevated anti-HER2 scFv expression, along with a substantial protein solubility enhancement at OD₆₀₀=0.6-0.8. In the LB medium, speeding up the shakers to 250 RPM led to a non-significant enhancement of anti-HER2 expression and a significant solubility improvement. The best medium for the optimum anti-HER2 scFv expression and solubility was the TB medium. MgCl₂ could not increase anti-HER2 expression or solubility. </span></span></span></span></span><br> <span style="font-size:10pt"><span new="" roman="" style="font-family:" times=""><b><span style="font-size:9.0pt"><span style="background:#2d7f8f"><span style="color:white">Conclusion: </span></span></span></b><b>&nbsp;</b><span style="font-size:9.0pt"><span style="color:black">The highest anti-HER2 scFv expression level was obtained when BL21-pET-22 (anti-HER2 scFv) was cultured in TB medium, induced at OD₆₀₀=2 and 37 &deg;C, and incubated at 250 RPM. The highest solubility was also observed in the TB medium, induced at OD₆₀₀=0.6-0.8 and 37 &deg;C, and incubated at 250 RPM.</span></span></span></span><br> &nbsp; Breast Neoplasms, Antibody Fragments, HER2 (Receptor, ErbB-2), Protein expression, Solubility 438 448 http://journal.zums.ac.ir/browse.php?a_code=A-10-5199-1&slc_lang=en&sid=1 Taranom Mobasheri taranom_1997@yahoo.com 5200319475328460089672 5200319475328460089672 No Ali Sharafi sharafi.a@gmail.com 5200319475328460089673 5200319475328460089673 No Elham Mohit el_mohit@yahoo.com 5200319475328460089674 5200319475328460089674 Yes