Fluorescence reader for microfluidic qPCR: faster, more sensitive and less expensive than most optical microscopes, it is a smart alternative for real-time fluorescence measurements of your on-chip qPCR signal. It offers a compact and cost-effective instrument compared to microscopes with a high numerical aperture, low magnification objective epifluorescence.
The FluoReader can be used to quantify on-chip qPCR signals in tiny chambers with an SNR that is more than 10 times higher than measurements done with most fluorescent microscopes equipped with a CCD camera. Thanks to its exceptional sensitivity, the FluoReader eliminates bleaching problems, reduces acquisition time and enhances the precision of acquisitions, allowing a more exact fitting of data and Cp calculation.
In this application note, we showcase the use of the FluoReader to perform qPCR.
The experiments shown in this application note have been realized only with Elveflow instruments and accessories. For any advice on your research project and experimental needs, do not hesitate to consult our team of specialists.
The complete qPCR system (figure on the left) consists of a heater controller (Cherry Biotech) commanding two heat exchangers. These two heat exchangers adjust the temperatures of heat-transfer liquids injected in the microfluidic chip in order to obtain the two desired qPCR temperatures for molecular samples. The switch between the two heat-transfer liquids within the microfluidic chip is achieved by a pressure controller (OB1 Mk3, Elveflow). The fluorescence detection is carried out by a FluoReader (figure on the right).
For qPCR assays, extracted and purified Bacillus atrophaeus (BG) bacteria DNA has been amplified. For each assays, 125 ng of initial DNA were used. 2 different qPCR master mix kits were used at a 1x concentration. An initial denaturation was conducted at 96°C and a temperature of 64°C was used for the annealing/elongation. The sample is contained in 1 mm x 1 mm x 100 µm transparent chamber.
The figure on the right describes the acquire signal during the qPCR. The lower panel shows the excitation light. Here, time-lapse detection is used to limit the light exposure of the fluorophore and the studied molecules.
400 µs light pulses are turned on every 1s. The light pulse consists of a binary signal of 0.5mW and 0 mW with of duty cycle 1/2. This osculating signal allows background removal by subtracting the signal detected when the light is ON to when the light is OFF. The signal is averaged with 0.4 s window and plotted in the upper panel.
The plot in the upper panel exhibits cycles of fluorescence signal which follows the thermalisation cycles. In fact the fluorescence of SYBR green changes inversely with the temperature. By following the upper part of the signal (when SYBR is attached to the DNA i.e. at low temperature), we see clearly an exponential increase of the fluorescence signal which shows the DNA amplification.
Have an interesting read on different detection systems used in microfluidics and the methodology behind them!
How can we help you?
Name*
Email*
Message
Newsletter subscription
We will answer within 24 hours
By filling in your info you accept that we use your data.
Do you want tips on how to best set up your microfluidic experiment? Do you need inspiration or a different angle to take on your specific problem? Well, we probably have an application note just for you, feel free to check them out!
Biofilm testing using a simple microfluidic chip channel for in situ observation of their development under flow conditions.
Microfluidics for microscopy imaging in plant biology allows to observe, in vivo, the biological response of plant roots to various stimuli.
This application note describes how co-culture of different cell types in separate but interconnected chambers is possible in a microfluidic platform
This application note explains how to study bacteria adaptation to stress and environmental changes such as antibiotics.
In this application note we describe how to set up medium recirculation by using microfluidic valves
In this application note we describe how to create a medium recirculation for dynamic cell culture with a microfluidic setup.
In this application note we describe how to stain cells for dynamic cell culture with different microfluidic setups.
In this application note we describe how to do cell perfusion for dynamic cell culture and a way to enable uni-directional recirculation of medium.
A simple guide to do dynamic cell culture by automating cell seeding in a microlfuidic chip
This application note proposes a microfluidic cardiac cell culture model (μCCCM) to recreate mechanical loading conditions observed in the native heart (in both normal and pathological conditions) by using an Elveflow OB1 pressure and flow controller.
In this application note, we will describe how to perform an automated and fast medium switch thanks to the Perfusion Pack.
Medium switch is widely used in cell biology. One application is the study of cell behavior under given flow conditions for different samples. In this tutorial, we walk you through the steps of a fast and stable medium switch using IBIDI© flow cells.
Prostate cancer is the second leading cause of cancer-related death for men. Circulating tumor cells (CTCs) are considered as a marker of early cancer diagnosis and disease severity. Their screening in blood is thus crucial to detect metastatic stage in cancer patients.
Until recently, microfluidic devices have been employed to support tissue-engineering experiments on basal lamina, vascular tissue, liver, bone, cartilage and neurons as well as organ-on-chips.
This application note presents how to perform cell culture on chip using the Cell and Biology Pack and the MicroSlides developed by ALine Inc.
Get a quote
Collaborations
Need customer support?
Serial Number of your product
Support Type AdviceHardware SupportSoftware Support
Subject*
I hereby agree that Elveflow uses my personal data Newsletter subscription
Message I hereby agree that Elveflow uses my personal data Newsletter subscription