How to keep monitoring iPSCs?

25 July 2023 1 5K Report

Hi everyone,I need to keep the iPSCs with no differentiation and scale them up in bioreactor for a week. Do you know how can I monitor iPSCs regularly without using FACS or antibodies? Thank you

Muhammad Jawad

If you need to monitor iPSCs without using FACS (Fluorescence-Activated Cell Sorting) or antibodies, there are alternative methods you can consider. Here are some non-invasive and label-free techniques that can be used to monitor iPSCs regularly:

Microscopic Observation: Regularly observe iPSCs under a phase-contrast or bright-field microscope. This allows you to monitor their morphology, growth, and confluency over time. You can identify characteristic iPSC colony features and ensure that they maintain an undifferentiated state.

Automated Cell Counters: Use automated cell counters, which can provide cell concentration and viability measurements without the need for labeling or fixation. These counters utilize trypan blue or other dyes that can distinguish live from dead cells based on their membrane integrity.

Metabolic Assays: Measure metabolites or waste products in the culture media to indirectly monitor the growth and health of iPSCs. Glucose consumption, lactate production, pH levels, and other metabolic parameters can give you insights into cell viability and proliferation rates.

Impedance-based Cell Monitoring: Impedance-based cell analyzers can be used to monitor changes in the electrical properties of the cell culture as iPSCs grow and proliferate. This method is label-free and non-invasive, allowing continuous monitoring over time.

RNA Expression Analysis: Regularly perform gene expression analysis using qRT-PCR (quantitative Reverse Transcription Polymerase Chain Reaction) to assess the expression of pluripotency markers (e.g., Oct4, Nanog, Sox2) and differentiation markers. Changes in marker expression can indicate shifts in the differentiation state.

Microfluidic Devices: Some microfluidic platforms allow continuous monitoring of iPSC cultures, providing information on cell growth, viability, and differentiation potential over time.

Bright-Field Time-Lapse Imaging: Employ time-lapse imaging with bright-field microscopy to capture real-time changes in iPSC colonies. This can help you identify any spontaneous differentiation events and track cell behavior over time.

Remember that combining multiple methods can provide a more comprehensive understanding of your iPSC culture's health and behavior. Choose the most suitable options based on your equipment availability and experimental requirements. Always ensure that any monitoring methods used are compatible with maintaining a sterile and controlled environment within the bioreactor.

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