Novel laser-based deposition of active protein thin films

This paper reports the deposition of active protein thin films by a novel laser-based approach termed matrix-assisted pulsed laser evaporation (MAPLE). We have deposited uniform 10 nm to nearly 1 mum thin films of insulin and horseradish peroxidase (HRP). We performed several experiments to characterize the chemical integrity of the deposited films. Matrix assisted laser desorption/ionization and liquid chromatography/electrospray ionization mass spectrometry experiments performed on MAPLE-deposited insulin films indicate that the laser-material interaction involved in this deposition technique does not modify the protein's mass. Fourier transform infrared spectroscopy experiments show that the chemical functionality and secondary structure of MAPLE-deposited HRP are nearly identical to those of the native protein. We also find that deposited HRP films retain their ability to catalyze the reduction of 3,3'-diaminobenzidine (DAB), suggesting that the active site of transferred proteins is unaffected by the MAPLE process. We also produced patterns and multilayers with feature sizes from 20 to 250 mum by depositing different biomaterials through a shadow mask. Patterns of physisorbed HRP were then protected from dissolution in an aqueous environment by a semipermeable polymer overlayer that was deposited in situ using pulsed laser deposition. This polymer membrane protects the protein pattern when it is exposed to DAB solution and enables the optical observation of HRP activity for spots as small as 2000 mum(2). These results demonstrate that MAPLE is a preferred technique for depositing active biomolecules for applications ranging from microfluidic sensor devices to gene and protein recognition microarrays.