Effects of cytoskeletal inhibitors and antibodies on Invaplex internalizationa

TreatmentSpecificity of treatmentInhibitor concn (μg/ml) or dilution% Inhibition (or enhancement)b
Cytochalasin BInhibits actin polymerization0.166.3*
ColchicineInhibits microtubule formation0.16.1
NMSPNegative antisera1:1004.6
HMSP 10Anti-IpaB/IpaC/LPS1:10090.6*
HMSP 8Anti-IpaB/IpaC/LPS1:10082.6*
MAb 2F1IpaB1040.6*
MAb 2G2IpaC1091.5*
MAb 2E8S. flexneri 2a LPS10(2.4)
  • a The inhibition of Invaplex internalization was investigated in BHK-21 monolayers by using cytoskeletal inhibitors or antibodies. Monolayers were treated with either cytochalasin B or colchicine, incubated with Invaplex 24 and fixed, and cells containing Invaplex were enumerated via fluorescence microscopy. Approximately 69% of the cells incubated with only Invaplex were positive for intracellular Invaplex. This served as the optimal uptake (0% inhibition) value to which all other treatments were compared.

  • For antibody treatments, serum from hyperimmune mice or MAbs were incubated with Invaplex before addition to BHK-21 monolayers. Treatment with Invaplex alone (positive control) resulted in 57% of the cells being positive for intracellular Invaplex. This served as the maximum uptake value (0% inhibition) to which all other antibody treatments were compared (see Materials and Methods). NA, not applicable; NMSP, normal mouse serum pool; HMSP, hyperimmune mouse serum pool.

  • b *, P ≤ 0.001 (chi-square analysis). When enhancement was found, the value is indicated in parentheses.