The mechanism of block of nicotinic acetylcholine receptor (nAChR) channels by purified antibodies from patients with myasthenia gravis (MG) was investigated by using an ultrafast system for solution exchange at outside-out patches. IgG of MG patients and controls was purified by using protein A-Sepharose columns. Probes from 9 seropositive MG patients and 3 seronegative MG patients were tested. As a preparation, cultured mouse myotubes expressing the embryonic-type nAChR channels were used. Twenty-millisecond pulses of 1.0 mM ACh were applied repetitively to outside-out patches. Outside-out patches were preexposed with IgG in concentrations between 0.1 and 200 mg/L during application of ACh pulses. The peak current amplitude was reduced to values between 6% and 71% of control for the 9 seropositive and 3 seronegative MG patients. The block was concentration dependent and fully reversible after washout of antibodies. Incubation with IgG from different control patients did not reduce the peak current amplitude. In addition, our findings with purified IgG from seronegative MG patients support the idea of the immunopathogenesis of this disorder and may allow the development of a diagnostic test for seronegative MG patients.