Monospecific, affinity-purified polyclonal antibodies reacting with the ami
no-terminal half of the mouse Toll-like receptor 4 (Tlr4) ectodomain failed
to block LPS effects and, to the contrary, were capable of inducing TNF sy
nthesis when applied to mouse macrophages and cross-linked with a secondary
antibody. This effect was observed with macrophages derived from C3H/HeN a
nd C57BL/10ScSn mice, but not with macrophages derived from C3H/HeJ or C57B
L/10ScCr mice, indicating a specific, Tlr4-dependent effect. Neither primar
y nor secondary antibody caused any response if administered in the absence
of the other reagent, nor was any response observed in cells from mice lac
king Tlr4, or bearing the Lps(d) mutation of Tlr4. These findings support s
everal conclusions. Tlr4, the essential transducer of LPS responses, may ac
t independently of LPS itself. LPS needs not be internalized, nor must it b
ind to a secondary target within the cell in order to exert its effect; rat
her, the receptor alone is required for initiation of a signal. The data ar
e consistent with the hypothesis that a conformational change in Tlr4 is re
quired for activation via this receptor, and reveal that the amino-terminal
half of the Tlr4 ectodomain is a target sufficient for antibody-mediated a
ctivation.