The purpose of this work is the optimization of a proteins’ purification on a silicon-based microfluidic Lab-on-a-Chip (LOC). The target procedure is composed by three distinct Native Polyacrylamide Gel Electrophoresis stages performed in the same matrix and in a 3D arrangement, by means of thin-film gold electrodes. This purification technique achieves different goals, fundamental to perform the subsequent electrochemical detection on-chip: analyte extraction from a pre-conditioned sample, analyte clean-up and pre-concentration. The optimization was focused on reducing losses of the system and time taken by the process, targeting Hemoglobin variants. The versatility of the system and of the employed techniques permits to extend results to other proteins and applications. The optimization required the set-up of a new acquisition workbench and the combination of experimental and analytical approaches.
L’obiettivo di questo lavoro è ottimizzare il sistema di purificazione di proteine di uno specifico Lab-On-a-Chip (LOC) microfluidico ed in silicio. La procedura in esame si compone di tre corse elettroforetiche native in gel di poliacrilamide (PAGE), eseguite in 3D all’interno di uno stesso gel, utilizzando elettrodi d’oro a film sottile. Questa tecnica di purificazione permette di preparare l’analita per la successiva analisi elettrochimica: oltre ad aumentarne il grado di purezza e pulizia, esso viene pre-concentrato ed estratto dalla matrice iniziale già pre-condizionata. L’ottimizzazione svolta si è concentrata sulla diminuzione delle perdite e del tempo di corsa delle varianti di emoglobina. La versatilità del sistema e delle tecniche impiegate permettono di estendere alcuni risultati ad altre proteine e ad altre applicazioni. Ottimizzare la purificazione ha richiesto la messa a punto di un nuovo sistema di acquisizione e la combinazione di approcci sperimentali e analitici.
Optimization of an electrophoretic proteins' purification system on a lab-on-a-chip
BIANCHI, GEA
2017/2018
Abstract
The purpose of this work is the optimization of a proteins’ purification on a silicon-based microfluidic Lab-on-a-Chip (LOC). The target procedure is composed by three distinct Native Polyacrylamide Gel Electrophoresis stages performed in the same matrix and in a 3D arrangement, by means of thin-film gold electrodes. This purification technique achieves different goals, fundamental to perform the subsequent electrochemical detection on-chip: analyte extraction from a pre-conditioned sample, analyte clean-up and pre-concentration. The optimization was focused on reducing losses of the system and time taken by the process, targeting Hemoglobin variants. The versatility of the system and of the employed techniques permits to extend results to other proteins and applications. The optimization required the set-up of a new acquisition workbench and the combination of experimental and analytical approaches.File | Dimensione | Formato | |
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https://hdl.handle.net/10589/141120