The interaction of hen egg white lysozyme with penicillin and a range of metal ions has been studied using microcalorimetry and related biophysical techniques. UV difference and fluorescence experiments on the binding of penicillin to lysozyme are ambiguous and reflect the variety of published observations, but there is no clear evidence that penicillin competes for example with N-acetylglucosamine (NAG) in the binding cleft. This is supported by ITC binding experiments which suggest that penicillin-G binds to a site on lysozyme remote from the active site. DSC studies are complicated by the thermal instability of penicillin-G in solution above 3