Quantification of the activity of biomolecules in microarrays obtained by direct laser transfer.
The direct-writing technique laser-induced forward transfer has been employed for the micro-array printing of liquid solutions of the enzyme horseradish peroxidase and the protein Titin on nitrocellulose solid surfaces. The effect of two UV laser pulse lengths, femtosecond and nanosecond has been studied in relation with maintaining the activity of the transferred biomolecules. The quantification of the active biomolecules after transfer has been carried out using Bradford assay, quantitative colorimetric enzymatic assay and fluorescence techniques. Spectrophotometric measurements of the HRP and the Titin activity as well as chromatogenic and fluorescence assay studies have revealed a connection between the properties of the deposited, biologically active biomolecules, the experimental conditions and the target composition. The bioassays have shown that up to 78% of the biomolecules remained active after femtosecond laser transfer, while this value reduced to 54% after nanosecond laser transfer. The addition of glycerol in a percentage up to 70% in the solution to be transferred has contributed to the stabilization of the micro-array patterns and the increase of their resolution.
Dinca
V
V
Institute of Electronic Structure and Laser (IESL), Foundation for Research and Technology Hellas (FORTH), P.O. Box 527, 711 10, Heraklion, Crete, Greece.
Ranella
A
A
Farsari
M
M
Kafetzopoulos
D
D
Dinescu
M
M
Popescu
A
A
Fotakis
C
C
D001681:Biological Assay; D064211:Connectin; D006735:Horseradish Peroxidase / Q000032:analysis* / Q000378:metabolism; D007834:Lasers; D008832:Microchemistry / Q000379:methods*; D009124:Muscle Proteins / Q000032:analysis* / Q000378:metabolism; D040081:Protein Array Analysis / Q000379:methods*; D011494:Protein Kinases / Q000032:analysis* / Q000378:metabolism; D013053:Spectrophotometry; D013499:Surface Properties
D064211:Connectin; D009124:Muscle Proteins; D006735:Horseradish Peroxidase; D011494:Protein Kinases