The qCamProto is an early hand crafted prototype of a qPCR / qLAMP device based on a PocketPCR and a webcam to read fluorescent signals. The prototype was built by Urs Gaudenz, GaudiLabs and first tested during the GlobalLAMP collaboration in Paris meetup in Paris . The results both for qPCR and qLAMP amplifications look very promising. The use of a webcam to read fluorescent can have advantages and disadvantages compared to sensing with a photodiode as discussed below.
qCamProto with a modified PocketPCR for heating and cooling. The heatblock was replaced by a 4 tube linear heatblock. The webcam is looking up to the heatblock, fixed with a 3D printed holder.
The webcam used is a 914519-3X0 (pulled form a HP laptop). The cam has a manual shutter (exposure) and manual gain option that is set to get a sensitive and stable reading.
Excitation and emission filters (foils) are installed in front of camera and LEDs
The signal curves are recorded using a prototype software written in Processing. The webcam image is captures over USB. The signal regions are masked and the brightness measured and plotted vs time. The blue LEDs for exciting the fluorescence is activated by the PocketPCR for a short periode of time the end of the cycle (to prevent bleaching). In this crude prototype the reading and excitation is not synchronized well and recovered data needed to be cleaned to compensate for that.
First results of qPCR amplification using Bacterial 16s rDNA primers for Bacterial identification. 3 positive tubes with a bacteria template and 1 negative control (bacteria was in LB while negative control was just water).
Pros and Cons of WebCam for fluorescent reading vs photodiode
In traditional commercial qPCR machines both, photodiodes and cameras (scientific grade CCD as sCMOS) are used to read the fluorescent signals. For low-cost devices photodiodes seem like the only choice considering the high cost of camera systems and optical setups. However the use of low-cost and readily available "webcam" consumer grade cameras without any additional optics might be good enough and open up new opportunities.
Possible advantages of camera setup:
- one camera for many tubes
- integrated amplifier, no need for extra amplifiers
- digital interface to computer or microcontroller
- adjustable exposure and gain
- reduced heating of the sensor due to optical distance to heatblock
- tubes can "be seen" on image
- active reading area can be choosen on image to improve signal (masking of reflections etc).
- image can potentially be used to detect presence of tubes / dirty tubes etc
- cost of good enough cameras as low as 1-2 USD
- maybe RGB separation can be used for something? (excitation and emission filters are still needed).
- signal of cameras not well know (limited information on data sheets)
- distortions and parallaxes
- complicated interface and drivers needed
- stable and precise mounting of camera on device
- (maybe) lower sensitivity
- no control over amplification stage
- long term availability of cameras not guaranteed
- light tight enclosure needed