Screening of Insect Cell Lines for the Production of Recombinant Proteins and Infectious Virus in the Baculovirus Expression System

Abstract

Eight cell lines derived from the insects Spodoptera frugiperda, Trichoplusia ni, Mamestra brassicae, and Estigmene acrea were evaluated for recombinant beta-galactosidase and infectious virus production following infection with the baculovirus Autographa californica multiple nuclear polyhedrosis virus (AcMNPV). Production was assessed on a specific (per cell and per microgram of uninfected cellular protein) and on a volumetric (per milliliter) basis. Cell density was found to be an important factor in comparing the cell lines due to a density-dependent inhibition of specific protein and virus production that appeared to result from cell-cell contact. After infection of cells at low-density specific beta-galactosidase production per cell would drop between 3- and 6-fold in five of the eight cell lines when plated on tissue culture plates at near-confluent and confluent cell densities. The cell lines Sf 21 and Sf 9 were least sensitive to cell density. After accounting for cell density effects and differences in cell size, two cell lines, BTI Tn 5B1-4 and BTI TnM, were identified that were superior to the other cell lines, including Sf 21 and Sf 9, in beta-galactosidase production. Optimal volumetric and specific beta-galactosidase production from Tn 5B1-4 and TnM cells was 2-fold and 5-fold higher, respectively, in both cell lines than the optimal production from Sf 9 or Sf 21 cells. The Tn 5B1-4 cell line also had the highest viability of all the cell lines at 3 days postinfection and could be adapted to serum-free media.(ABSTRACT TRUNCATED AT 250 WORDS)

Antigenic and immunogenic analysis of group A and group B respiratory syncytial virus G proteins expressed from recombinant baculoviruses

“…Spodopterafrugiperda (Sf9) cells were maintained in Grace's medium (Gibco/BRL) supplemented with 10 % FCS and PKS. A cell line derived from Triehophtsia ni eggs, Tn 5Bl•, (Wickham et al, 1992) was obtained commercially (High 5 cells; Invitrogen) and maintained in Excell 401 medium (JRH Biosciences) with PKS.…”

A new rapid amplification of cDNA ends method for extremely guanine plus cytosine-rich genes

“…mRNA was isolated either from second instar Trichoplusia ni larvae (Chesapeake-PERL, Savage, MD) or from the Trichoplusia ni cell line, BTI-Tn5B1-4 (High Five®; [14]). Briefly, total RNA was isolated from one million BTI-Tn5B1-4 cells or from one second instar larva using 1 mL of Tri-Reagent® (MRC, Cincinnati, OH) according to the manufacturer's instructions.…”

Transfection of Spodoptera frugiperda Insect Cell and Production of Recombinant Protein

“…İki hücre hattı da transfeksiyon, saflaştırma, yüksek titreye sahip stoklama ve rekombinant protein üretimi için uygundur. Bununla birlikte Sf21 hücre hattı monolayer kültür ile üretimde Sf9 hücresine oranla daha hassas olması nedeniyle pasajlamada canlı hücre kayıpları meydana gelmektedir [11]. Spodoptera frugiperda'nın pupal evredeki yumurtalık dokularının Autographa californica çekirdek polihedrosis virüsü (AcMNP) ile enfeksiyona duyarlılığının yüksek olması nedeni ile tüm baculoviral ekspresyon vektörleri ile kullanılabilmektedir.…”