Improving the expression of recombinant soluble HIV Envelope glycoproteins using pseudo-stable transient transfection.
The Envelope glycoprotein (Env) of the human immunodeficiency virus (HIV) is the target of neutralizing antibodies (NAbs). So far, HIV Env-derived immunogens have not been able to elicit broad neutralizing antibody responses against primary isolates. Identifying conditions that will permit the efficient production of different soluble HIV Env proteins will facilitate a high throughput comparative analysis of the immunogenicity of diverse Env constructs, potentially identifying Env forms that are more conducive to the elicitation of anti-HIV NAbs. Here we compared different cell types, transfection reagents, transfection conditions and different DNA expression vectors on soluble HIV Envelope expression levels. We identified optimal expression conditions and developed a protocol to streamline and maximize production of diverse HIV Env constructs. Using this optimized platform, milligram quantities of purified soluble HIV Env trimer can be routinely achieved in a rapid and cost-effective manner.
Seattle Biomedical Research Institute, Seattle, WA 98109, United States.
George
G
Sellhorn
Zachary
Z
Caldwell
Christine
C
Mineart
Leonidas
L
Stamatatos
D015153:Blotting, Western; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D006678:HIV / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D014162:Transfection / Q000379:methods*; D054299:env Gene Products, Human Immunodeficiency Virus / Q000235:genetics / Q000378:metabolism*
D011994:Recombinant Proteins; D054299:env Gene Products, Human Immunodeficiency Virus