Operation of Laboratory Photobioreactors with Online Growth Measurements and Customizable Light Regimes
October 28th, 2021
This publication describes the design of laboratory photobioreactors (PBRs) with customizable light regimes. The growth of cyanobacteria or microalgae, using bicarbonate as their carbon source, is monitored continuously by measuring volumetric oxygen production. These PBRs facilitate rapid, replicated laboratory growth comparisons with little user intervention during experiments.
This protocol enables replicated microalgae experiments with little user input. It is designed for researchers looking to upgrade from manually monitored to bench top flasks without purchasing technical and expensive commercial photo bio reactors. This photo bio reactor enables light ratio and flexibility, continuous growth monitoring, and triplicate replication.
To begin, open the data acquisition software. Fill in the configuration page and assign calibration files to their respective sensors. Under the directory file name, select the corresponding digital input module port number folder.
Click on the current folder and repeat for all ports. Then click on okay to move on to the logging page. Next, fill the glass bottles to the desired volume with the cultivation medium and centrifuge the stock culture in three balanced 50 milliliter tubes to yield three pellets.
Add one pellet to each bottle using a serological pipette and fresh medium. Then drop a 25 by eight millimeter magnetic stirrer into each bottle before sealing the bottle opening with a rubber stopper and threaded aperture screw cap. If any optional sampling ports are installed, close the valves.
After locating the end of each gas line inside the PVC pipe, attach a needle to the valve's mail lure port. Connect each bottle to its gas sensor by piercing each rubber stopper with the corresponding needle. Then, launch each gas sensor individually by checking the tick box on the left side of the screen, clicking on start, and inputting a file name.
Click okay and repeat for all sensors. Next, switch on the photo bio reactor and ensure that the digital multiplex lighting controller is plugged into a power supply. Using the button on the digital multiplex controller, switch between the helper scene and the customized light scene to ensure the device is functioning correctly.
For bottle sampling, prepare an additional 500 milliliters of the fresh medium before commencing the experiment. To collect the sample, close the valve on the gas line and connect a 10 milliliter syringe to the sampling port valve. After connecting the syringe, open the sampling port valve and withdraw eight milliliters of culture.
Then close the sampling port valve and disconnect the syringe. Next, connect to syringe containing eight milliliters of fresh medium to the sampling port valve. Then open the valve and inject the fresh medium.
Finally, close the sampling port valve and disconnect the syringe. To export the data, select file and offline data. After selecting all relevant log files, export the data to spreadsheet software and save.
A successful experiment is characterized by a closely replicated diurnal pattern of oxygen production. The oxygen flow rate acts as a proxy for a culture's growth rate. During illuminated hours, gas production steadily increases and during non illuminated hours, gas production stops.
The total volume of oxygen produced over four days ranged from 316 milliliters in treatment A to 902 milliliters in treatment C.In the 300 micro mole per meter squared per second light regime, the temperature increased by a maximum of 1.4 degrees Celsius during the day. Culture temperatures returned to baseline overnight. Despite differing in their initial biomass concentrations, treatments B and C generated the same amount of total biomass causing identical shifts in medium pH.
The online oxygen flow rate data revealed that each treatment had varying daily growth rates, which was also reflected in the twice daily pH measurements. On day one, treatment B's growth rate was lower than that of treatment C.Whereas by day three, treatment B's growth rates surpassed that of treatment C.Oxygen flow rate data indicated that the highest growth occurred on day three in treatment B.Growth estimates based on total measured oxygen were expected to slightly underestimate biomass growth which was valid for five out of seven examples. Two examples overestimated growth.
This may have been influenced by experiments being determined at the end of the night. Substantial nighttime respiration enhances oxygen consumption, generates a bottle head space vacuum, sucking in pack with liquid from gas sensors. If respiration may be high, we opening the head space overnight.
Overview
This publication outlines a protocol for designing laboratory photobioreactors (PBRs) suitable for microalgae experiments. It highlights the continuous monitoring of oxygen production as a proxy for growth analysis, aimed at minimizing user intervention. The PBRs allow for customizable light conditions and facilitate rapid growth comparisons.
Key Study Components
Research Area
- Cyanobacteria and microalgae growth
- Photobioreactor design
- Carbon source utilization
Background
- Need for efficient growth monitoring methods
- Limitations of manual cultivation techniques
- Cost-effective alternatives to commercial photobioreactors
Methods Used
- Assembly and configuration of PBRs
- Cyanobacteria or microalgae as model organisms
- Volumetric oxygen production measurement
Main Results
- Successful monitoring of oxygen flow rates correlated with growth rates
- Observations of diurnal patterns in oxygen production
- Findings suggest variations in growth dynamics among different treatments
Conclusions
- The study demonstrates a viable protocol for flexible and low-intervention growth monitoring of microalgae.
- It contributes to the advancement of methods to explore algal biology and optimize cultivation techniques.
Frequently Asked Questions
Executive Industry Relevance
This protocol enables scalable, replicated microalgae cultivation with automated growth monitoring and programmable light regimes, addressing the need for high-resolution phenotypic data in early-stage target validation. By correlating oxygen flux with biomass accumulation, it provides quantitative, time-resolved outputs that support mechanistic de-risking of photosynthetic pathway engineering. The system’s low-cost, modular design facilitates enterprise reuse across discovery biology and assay development workflows.
Strategic Applications in Biopharma R&D
Early Discovery & Target Validation
- Scientific Value: Enables interrogation of photosynthetic efficiency and carbon fixation pathways under controlled light conditions.
- Operational Value: Supports triplicate replication for statistical confidence in target phenotype assessment.
- Predictive Value: Generates growth rate proxies that help prioritize strains with desired metabolic traits.
Screening & Assay Development
- Assay Readiness: Provides continuous, high-resolution oxygen production data as a quantitative biomarker of culture growth.
- Scalability: Vertical PVC pipe design allows multiplexed testing of light regimes within a small footprint.
- Reproducibility: Automated logging minimizes user intervention and ensures consistent data collection across experimental runs.
Translational & Preclinical Research
- Disease Relevance: Supports screening of microalgae strains for recombinant protein or bioactive compound production.
- Translational Continuity: Enables progression from strain characterization to biomass yield optimization under defined conditions.
- Risk-Adjusted Advancement: Oxygen-based growth metrics allow objective comparison of strain performance before scale-up.
Pipeline & Workflow Integration
The method integrates into early discovery by enabling hypothesis testing of genetic modifications affecting photosynthetic output, feeds into screening via standardized growth assays, and supports translational research through scalable biomass production under tunable light.
- Discovery Biology: Facilitates pathway clarification by linking light regime adjustments to measurable changes in oxygen evolution.
- Screening: Delivers assay-ready, time-series growth data for comparing strain responses to environmental variables.
- Analytics: Oxygen flow rate measurements serve as a proxy for biomass accumulation, enabling quantitative comparison across treatments.
- Translational Research: Supports scale-up decisions by providing reproducible growth curves under industrially relevant light conditions.
- Enterprise Reuse: Modular PBR design allows replication across labs for standardized strain evaluation campaigns.
Operational & Enterprise Impact
- Scientific Value: Reduces mechanistic ambiguity in strain selection by providing direct, real-time readouts of photosynthetic activity.
- Operational Value: Ensures standardization and reproducibility through automated sensor logging and environmental control.
- Strategic Value: Improves go/no-go decisions by enabling data-driven prioritization of high-performing strains.
- Portfolio Impact: Supports risk-adjusted advancement by correlating early growth metrics with potential biomass yield.
Implementation Considerations
- Requires expertise in microbial culture maintenance and sensor calibration.
- Needs data acquisition hardware, multiplex lighting controller, and volumetric gas sensors.
- Demands cross-team standardization of light programming and gas sampling protocols.
- Adaptation to different model systems may require adjustments to gas exchange and mixing configurations.
- Practical limitations include headspace vacuum risks during high respiration periods, necessitating periodic venting as noted in the source.
