Pseudotype vectors are promising for gene transfer in many gene therapy approaches, however low vector concentration in batch cultures and high temperature-dependent decay limit sufficiently large-scale production. The ability of commercial ceramic asymmetric ultrafiltration membranes with a cut-off of 20 kDa to purify retroviral pseudotype vectors derived from the murine leukemia virus (MLV) carrying the HIV-1 envelope protein MLV (HIV-1) was studied. Experimental study was carried out in order to analyse the impact of inline tangential flow filtration (TFF) combined with different harvest temperatures in continuous production mode compared to batch filtration. Retroviral pseudotype vectors were produced using a 200 ml fixed bed reactor for high cell density cultivation on macro porous carriers up to 400 hours. By tangential flow ultrafiltration combined with cooling down the harvest supernatant to 4 degrees C, the vector concentration was increased 12-fold with an average recovery of 78.7% of the initial infective capacity.