Vertical microbubble column-A photonic lab-on-chip for cultivation and online analysis of yeast cell cultures

Abstract

This paper presents a vertically positioned microfluidic system made of poly(dimethylsiloxane) (PDMS) and glass, which can be applied as a microbubble column (μBC) for biotechnological screening in suspension. In this μBC, microbubbles are produced in a cultivation chamber through an integrated nozzle structure. Thus, homogeneous suspension of biomass is achieved in the cultivation chamber without requiring additional mixing elements. Moreover, blockage due to produced carbon dioxide by the microorganisms-a problem predominant in common, horizontally positioned microbioreactors (MBRs)-is avoided, as the gas bubbles are released by buoyancy at the upper part of the microsystem. The patterned PDMS layer is based on an optimized two-lithographic process. Since the naturally hydrophobic PDMS causes problems for the sufficient production of microbubbles, a method based on polyelectrolyte multilayers is applied in order to allow continuous hydrophilization of the already bonded PDMS-glass-system. The μBC comprises various microelements, including stabilization of temperature, control of continuous bubble formation, and two optical configurations for measurement of optical density with two different sensitivities. In addition, the simple and robust application and handling of the μBC is achieved via a custom-made modular plug-in adapter. To validate the scalability from laboratory scale to microscale, and thus to demonstrate the successful application of the μBC as a screening instrument, a batch cultivation of Saccharomyces cerevisiae is performed in the μBC and compared to shake flask cultivation. Monitoring of the biomass growth in the μBC with the integrated online analytics resulted in a specific growth rate of 0.32 h(-1), which is almost identical to the one achieved in the shake flask cultivation (0.31 h(-1)). Therefore, the validity of the μBC as an alternative screening tool compared to other conventional laboratory scale systems in bioprocess development is proven. In addition, vertically positioned microbioreactors show high potential in comparison to conventional screening tools, since they allow for high density of integrated online analytics and therefore minimize time and cost for screening and guarantee improved control and analysis of cultivation parameters.

Figure 1

Schematics of the μBC: (a) patterned PDMS layer, (b) microstructured glass bottoms with gold mirror for optical detection or with microheater, (c) PDMS-glass-microchip, (d) zoom in of implemented fluidic interface, (e) zoom in of the out-of-plane lens design that is patterned in PDMS to optically monitor bubble formation, (f) zoom in of implemented optical elements for OD monitoring in x-axis (in-plane).

Figure 2

Image of the fabricated μBC: (a) with gold mirrors, (b) with microheater. (c) Photograph of one of the fluidic interfaces.

Figure 3

Schematics and photographs of the adapter module made of polycarbonate including the chip carrier for the μBC microchip and the fluidic (including fit borings and fit bolts), electrical, and optical connectors. The two different optical bridge connectors for out-of-plane OD monitoring comprise either (1) an LED and photodetector (used for bubble formation monitoring) or (2) two optical fibers.

Figure 4

Voltage signal of the photodetector versus time demonstrating (a) unstable and (b) stable bubble formation.

Figure 5

Absorbance versus PMMA particle concentration of the OD measurement (a) out-of-plane and (b) in-plane.

Figure 6

Schematic setup of the cultivation procedure in the μBC.

Figure 7

OD versus cultivation time in the μBC and the shake flask.