Difference between revisions of "Team:Grenoble-Alpes/temperature module"

TEMPERATURE MODULE

• PROJECT
  • BIOLOGY
  • ENGINEERING
    • PIPETTING MODULE
    • FLUORESCENCE MODULE
    • TEMPERATURE MODULE
    • PURIFICATION MODULE
  • DEMONSTRATE

In the automated system, many steps require to heat or cool biological samples. Typically, the biological transformation is a good example: the samples containing the bacteria and the DNA need to be heated and cooled for precise durations at a constant temperature to realize a heat shock (a classical transformation goes from 0°C for 30 minutes to 42°C for 1 minute and back at 0°C for 2 minutes). Hence, this module has to be carefully considered because it is a key point to the success of any biological manipulation. In addition, because our system will be fully automated and nobody will be able to check the inside of the machine while it works, a reliable and robust module is required so that each time it is used, repeatable results are obtained.

In the following figure, the different biological steps are summarized and their required temperatures are given.

By design, each biological step will be contained in a module with a given temperature. This allows us to make temperature shocks by pipetting the liquid inside an Eppendorf tube from one place with a given temperature (0°C for instance) to another place in the machine with another temperature (42°C this time).

Design of the module

An Infra-Red temperature sensor placed on top of the Eppendorf tube can continuously sense the temperature of the solution from a distance (without disturbing or contaminating it) and send it to an Arduino.

A thermal conductor, placed around the tube and in contact with the heating/cooling device, eases and homogenizes the heat/cold transfer between the Eppendorf tube and the device producing heat/cold (to better homogenize the solution, an electronic pipette can also flush the solution).

The device in charge of heating/cooling the solution is placed under the Eppendorf tube. It is activated with the help of an Arduino and a MOS transistor. A goal temperature is programmed in the Arduino and the heating/cooling device is switched on/off to be kept at this goal temperature (this way of programming the heating/cooling device powering is called an “all or nothing regulation”).

Temperature tests

In the following part, measurements of the temperature inside an Eppendorf tube placed in the modules were made to characterize them.

The cold module

The temperature inside a tube placed in the module (red curve) was measured and compared to the evolution inside of an ice tank (green curve).

During the test, we were only able to make the module go down to only +11°C. The main explanation would be that 2 Peltiers modules are not enough to cool down eight Eppendorf tubes at the same time. The Peltier modules are still capable of cooling the tubes from 27°C to 11°C (16°C difference).

However, the slope is quite similar for both curves. The 8 solutions still undergo a rapid change of temperature that can be considered as a temperature shock and might be enough for the success of a bacterial transformation. The effectiveness of it will have to be further tested. Also, because obviously this system will never reach 4°C (the goal temperature at which it should be powered-off by the all or nothing loop), the system is continuously powered so it doesn’t risk a degradation and the temperature will stay constant on its own. No variations to potentially disturb the bacteria.

        The other interesting information here is that it takes 10 minutes (600 seconds) for the module to get to its lower temperature. Hence, before any manipulation, the user will have to power-up the machine at least 10 minutes before using it.

The heating module

The heating module works in the same way as the cooling module but the temperature is taken inside an empty Eppendorf tube so that the whole environment is at the goal temperature and not the liquid inside of the tube.

A goal temperature is set (42°C here) and the evolution was plotted in the previous figure. It takes about 15 minutes for the module to go to 42°C and once it has reached the goal temperature, the power turns off for a little bit of time, causing the temperature to drop again to 37°C. This shows an inaccuracy of the temperature of about 5°C that could be problematic if we heated the solution for a longer time at 42°C (which is not the case because in a bacterial transformation, the tubes are heated for about one minute).

Moreover, when we put an Eppendorf tube filled with liquid inside the module, the IR sensor will measure that the solution is under 42°C and power-on the module so that the temperature will immediately go up. In the end, when the tube is put inside of the module, the temperature is always between 40°C and 42°C, which is a better accuracy.

In the case of a goal temperature of 70°C, the heating resistor powered by a 12V supply is only able to heat-up the whole aluminum block to 70°C +/- 2°C. Hence, the control loop will rarely be triggered and the temperature will stay constant on its own at 70°C.

Moreover, it is interesting to see that for any step requiring a temperature àf 70°C, the module needs to be started 7 minutes beforehand to get to the right temperature.

How to build the modules ?

LEARN MORE

About the choice of components producing heat/cold

The following paragraph will focus on the choices of components depending on their characteristics.

The Peltier module

It is often much easier to generate heat than a cold source. However, after some research, we found out about the Peltier module (see figure below) [1].

A Peltier module is made of two different semiconductors linked by two junctions. When a current is applied to one of the semiconductors, heat is liberated at one junction and cold at the other junction. To sum up, when we apply a current to a Peltier module, one face becomes very hot and the other very cold [3].

However, due to heat diffusion from hot areas to cold areas, after some time, the whole Peltier module becomes hot. This is why when the cold face is the one used, a ventilation cooling system has to be placed on the hot face of the Peltier module so that as soon as heat is produced on this face, it is ventilated outside of the system. The better the ventilating system is, the more cold is produced by the Peltier.

After comparing several Peltier modules performances, I chose a two-stages Peltier module (see figure 12).

The main advantages of this Peltier module is that it produces a lot of cold without requiring a huge cooling system. It can work with a 12 Volts (V) power supply and it does not require more than 3A unlike one-stage Peltier modules who often need up to 6 Ampers (A) to work, which is very difficult to supply when several devices are plugged on the same power supply. When other types of Peltier modules were tested, the cold surface would also get hot very easily without a big radiator similar to the 13th figure to evacuate the generated heat (about 20 centimeters long).

With the idea of an automated machine in mind, this type of radiator was too voluminous to be integrated into the system. We preferred a much smaller and practical cooling system represented in the 14th figure. This system was obviously less effective than the previous one but it was enough for our use:

Heating resistor

To heat-up a solution, a heating resistor [5] was used. This kind of device converts electrical energy into heat when powered on (12V supply).

Among equivalent devices, the resistor used was chosen for its shape: it is very thin and plane. The copper tube containing the Eppendorf tube was placed on top of the resistor so that the solution could be heated from the bottom. However, this resistor was only able to reach 74°C so it was used preferably for the extraction, incubation and transformation steps where the temperature is below 74°C.

Moreover, because of a lack of communication between the microbiologists of the team and the engineers, I learned very late in the project that we needed to go up to 91°C for the hybridization step and I had to find a solution in less than a week.

A Peltier module to heat

At first, it seemed impossible to find a small heating resistor powered with a 12V supply that could heat a solution up to 91°C within a respectable delay. Then I remembered of the one-stage Peltier modules I disregarded because the cold surface would get hot too easily. But after testing heating a solution with the hot side of a Peltier module, I was able to raise the water temperature up to 91°C in about 15 minutes.

About the electronic components choice

The MOS transistor

In the very first electronic circuit, a classical NPN transistor was used but it was getting hot very rapidly even when we put a heat sink behind it. The main reason is that a high of current is going through the transistor to power on the Peltier module or the heating resistor (up to 1A) and it is partially dissipated by heat. But, too much heat will damage the transistor.

According to the datasheet of a NPN transistor [6] (TIP200 for instance), the maximum intensity that a regular NPN transistor can endure is 5A. This explains why around 3A, the NPN transistor gets hot.

However, if a MOS transistor [7] is used (a BUZ100 for example), the transistor can endure up to 60A. Hence, when 3A is going through the transistor, it does not get hot.

The temperature sensor

We chose to use an Infra-Red temperature sensor [8] because it can measure the temperature without disturbing or contaminating the liquid inside of the tube.

It can measure temperatures from -40°C to 100°C, which covers the range of temperature we need to sense (from 0°C to 91°C). However, we have to consider the distance at which it is placed from the tube. The farthest the sensor is, the more the IR beam is self-diffracted and become large and unfocused. Consequently, the temperature given by the sensor loses precision because it is taken on a larger surface (the resulting temperature value is a mean of the beam surface). Considering that the Eppendorf tube has a diameter of 8 millimeters, we chose a sensor with a diffraction of 10°/cm and we place the sensor 1 centimeters away from the tube. Hence, the beam diameter is 0.9 mm wide, which is largely inferior to the diameter of the tube.

With this sensor, we are sure that the temperature we get is from the solution inside of the tube and not from the plastic of the tube for example.