Water circulator

Is there any reason why I couldn't just use my lab's temperature controlled water bath in lieu of a circulator?

Comments

  • Please see my comment here: http://forum.claritytechniques.org/discussion/10/bubbles-in-the-tissue#latest

    Having the clearing solution circulate through the chamber is necessary to prevent bubble accumulation in your sample. In addition, it prevents by-product buildup on the electrodes which will alter the composition of the clearing solution (usually indicated by a drop in pH). If the pH is not maintained, clearing will not occur and your sample could become damaged.
  • achungachung Posts: 7
    edited June 2013
    I understand...I was thinking I could use something like the pumps used for perfusions to circulate the solution. Or even a fish pump. Would that work?
  • rajuraju Posts: 12
    I think it should work fine, as long as you have a reasonably fast way to cool down the clearing solution (to dissipate heat generated in etc). Not sure how fast "temperature controlled water bath" can control the temperature?
  • https://docs.google.com/file/d/0B6RE82086JGFT1gxYlZldnE4Vlk/edit?usp=sharing

    https://docs.google.com/file/d/0B6RE82086JGFRzBDa1U2SFQ4Y3M/edit?usp=sharing

    https://docs.google.com/file/d/0B6RE82086JGFYTR1Z3hwRldkQ1U/edit?usp=sharing

    @achung this is my setup, basically what you described. Fishtank circulator and waterbath. Clears tissue nicely, and the fishtank circulator is more than enough to clear the bubbles out of the tissue.

    @raju. As long as your buffer filter and circulator are both left in the bath, the water bath actually controls the temperature of the circulating solution very well.
  • jberryjberry Posts: 5
    What is the model of your circulator? What is the flowrate?
  • NamasraviNamasravi Posts: 1
    @joelsosiene I am also planning to use fisk tank pump. Could u kindly let me know the flowrate of pump and the make??? and how do u adjust the flowrate????
  • I use a Zoomed TCN30, which circulates at 5 liters per minute. I found that the circulating speed was a little fast, so I included a wall within my chamber which breaks up the turbulence and decreases effective flow. Remember I am using a non-standard design.
  • SVSUNEUROSVSUNEURO Posts: 12
    @joelrosiene Any thoughts on possibly daisy chaining your chambers? If so, how many do you think would be possible?
  • FabioFabio Posts: 62
    this is possible and people do it, there are comments on this in the forum
  • I have daisy chained 3 chambers in a row, using the same circulator and only one power supply. Limiting factors are essentially just the amperage cap on your PSU, and I guess the decrease in flow rate that daisy chaining the chambers produces.
  • I'm using a Cole-Parmer EW-77910-30 peristaltic pump (~$900 new) set at about 150 ml/min to pump buffer from a 2L bottle in a 37C water bath into the chamber. The chamber then drains back into the bottle. This has been keeping the temp of the chamber outflow at ~38-39C and easily removing all the bubbles. This peristaltic pump/head combo can pump up to 500 ml/min. I change my buffer daily.
  • GuptaJGuptaJ Posts: 4
    @suzschindler Is the pump tubing (Cole-Parmer EW-77910-30 peristaltic pump ) sturdy enough to be run for 7-8 days continuously. For how many days are you doing the ETC run?
    I actually used a regular perfusion pump we use in lab for short time perfusion and unfortunately the tubing broke after a day of continuous run and the brain got cooked.
  • XWongXWong Posts: 3
    @GuptaJ - I also use a similar perfusion pump for my ETC set-up (flow rate of approx 500mL/min, 20V, 37C). The tubing was damaged after 5 days of continuous run, and at some point there was just no more clearing solution in circulation (leaked all over the lab bench and floor during the weekend). But what did you mean by "cooked brain". I didn't notice any obvious change in the texture/color of my mouse brain after this incident...
  • GuptaJGuptaJ Posts: 4
    @XWong I was running the ETC set up (8L/hr, 20V, 37C) with perfusion pump which pumped buffer from a 2L bottle and finally drains the clearing solution back in the bottle (maintained at 37 C). The setup was quite similar to what @suzschindler used except for the perfusion pump. I was using Fisher Scientific Mini pump variable flow (medium/high flow model). After a day, because of the constant run and friction produced (on the tubing around the rotating wheel), the tubing of the pump broke. The clearing solution was all over the lab bench. Meanwhile, the brain sample was still in the ETC chamber and the electrodes were connected to the electric supply. But because the pump failed, there was no more clearing solution in the chamber. and in the constant electric field , my sample burnt and reduced to a small brown lump. (even though the brain was well away from the electrodes and safely enclosed in the falcon strainer)
  • To those using perfusion pumps - I have had good results with C-flex Masterflex tubing from Cole-Parmer. I ran ETC continuously for 6 days with no leaks.
  • GuptaJGuptaJ Posts: 4
    @liz_kritzer Which Pump are you using? The pump that I'm using would not support the tubing you suggested.
  • @GuptaJ I am using a Cole-Parmer Masterflex pump #7553-10 with pumphead 7017-20. It's a pretty old pump that we had sitting around in the lab.
  • GuptaJGuptaJ Posts: 4
    Thanks a lot @liz_kritzer
  • For people using a peristaltic pump, I recommend Excelprene TPE tubing. I use it anytime I need to run something through a pump and it has always worked really well. Has really high fatigue resistance and meets FDA, 3-A and NSF criteria with a temp range up to 275°F. Available at pretty good prices in 25 ft rolls from U.S. Plastic:

    http://www.usplastic.com/catalog/item.aspx?itemid=85322&catid=571
  • I move the tubing a few inches every day to prevent the rollers from rubbing through. Doing this, I never have the tubing break.
  • achungachung Posts: 7
    @joelrosiene do you have the clearing solution in the water bath as well, or do you have just water in the bath, and then the solution just circulating?
  • I have a 2 L bottle filled to 1.4 L in a water bath. I didn't really want to ruin/contaminate the water bath with the buffer. The solution just runs in a loop--into and out of the same bottle.
  • @achung Many apologies for such a belated reply - the solutions in the bath and the circulator do not mix, therefore it is not necessary to have clearing solution in the bath. It's just tap water.
  • Hey there,

    we are having some difficulties performing ETC. The biggest problem seems to be caused by our pump which we use to circulate the Clearing Solution. It cannot provide a sufficient flow rate and temperature. Therefore I was looking for a reasonable and affordable replacement.

    Maybe someone can take a look at the link below and give me some advice.

    http://www.julabo.com/us/products/heating-circulators/heating-circulators-with-open-bath/ed-5-heating-circulator-with-open-bath
  • ClarityNLClarityNL Posts: 14
    edited April 2014
    @Anatomy_Greifswald‌ : You can also use an ordinary water pump destined as aquarium pump. Our water circulation is regulated that way, with sufficient flow at low cost. Put the tank or bottle of clearing solution in a waterbath and its temperature can be controlled as well.
  • yetitiyetiti Posts: 7
    @Anatomy_Greifswald‌

    You should look for a circulator with cooling capacity while running ETC. The link you gave seems to only have heating ability.

    Hey there,

    we are having some difficulties performing ETC. The biggest problem seems to be caused by our pump which we use to circulate the Clearing Solution. It cannot provide a sufficient flow rate and temperature. Therefore I was looking for a reasonable and affordable replacement.

    Maybe someone can take a look at the link below and give me some advice.

    http://www.julabo.com/us/products/heating-circulators/heating-circulators-with-open-bath/ed-5-heating-circulator-with-open-bath

  • @yetiti‌

    Yes, the water circulator mentioned in the materials has cooling capacity but to me it seems like this was never really used. I just read that the temperature was set to 40-50°C with a recommended 37°C. It depends on the product which range of temperature it can provide but the one used by the D-Lab has the capacity of cooling below 0°C. Therefore it seems like this wide range was never really used or am I getting it wrong?

    Nevertheless the biggest problem with most water circulators seems to be setting of flow rates. Most products only provide 2-4 Setup levels which all seem to be too high because these levels are programmed as a ratio of the maximum flow rate.

    I spoke to a staff member of Lauda and he suggested a product which has the same configurations to the one used in Stanford without the great cooling range. Thus, it is much cheaper. He said it will work just fine at the temperatures and the flow rate we are trying to provide.

    Here a link to the Lauda product:

    http://www.lauda.de/hosting/lauda/website_en.nsf/urlnames/product?OpenDocument&ProductID=090429-20185-CC






  • @yetiti‌


    I was putting a lot of thought into your comment with the cooling capacity. I guess you wanted to point out the possibility of overheating in the chamber, didn't you?

    In our experience with a different set-up including a different pump the heat production in the ETC chamber seemed at least not relevant. In my opinion the water circulator posted in the link above has enough accuracy in temperature. Although it might take a while until it compensates an overheating in the chamber because it misses the cooling capacity.
    Additionally I think that a flow rate in the range of 1-3 L per minute is such a big flow compared to the volume of the chamber that it will cool the overheat as well.
  • FabioFabio Posts: 62
    I have a Lauda A6 (no cooling) at 20 - 30 V the temperature is stable
  • daj1u06daj1u06 Posts: 2
    Any consensus yet on flow rates? Posts on the forum seem to range from 150ml/min to theoretical 10l/min - given the chamber volumes all of these seem quite high - only asking because I'm about to setup a system and am still trying to work out the best pumping solution (maybe a peristaltic detergent dosing pump from a commercial dishwasher as these are way cheaper than lab models - anyone using this setup?). I would guess that you could get bad foaming with an impeller driven aquarium pump? Thanks, DAJ
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