Stir Bars

  1. Drop-seq: Single-Cell RNA-Seq Protocol

    The new Single-Cell RNA-Sequencing protocol called “Drop-Seq” uses V&P Scientific’s Magnetic Tumble Stirrer {VP 710D2}, Support Stand {VP 710D2-4}, and Magnetic Stir Discs for stirring in a syringe on a syringe pump.

     A very interesting paper was recently published in Cell titled, “Highly Parallel Genome-Wide Expression Profiling of Individual Cells Using Nanoliter Droplets,” by Evan Macosko et al in the Steve McCarroll lab at Harvard.

     As a former bench scientist who has done her share of gene expression analyses the really old fashioned way, from large numbers of cells of singular origin, like a culture flask of yeast, I can attest that this Drop-seq method is a powerful system these researchers have developed to study complex mixtures of cells at the single cell level.

    Read their article to see how the Drop-seq method can be used in your own research. 

    I will explain the use of V&P Magnetic Tumble Stirrer in the Drop-seq protocol to ensure proper setup & use for fellow researchers.

    I would follow what the researchers have outlined in their protocol for the mixing of the beads. However, I would like to add some details to their setup description. They have a nice diagram of the relative positions of the syringe pumps and the microscope, but it does not include the position of the VP 710D2 relative to the syringe pump it is stirring. They do describe putting the syringe pump in a vertical position such as this:

    But what they don’t mention is the VP 710D2-4 Support Stand, which was designed for use when mixing in syringes on syringe pumps. The Stand positions the VP 710D2 above the syringe  to allow for more effective stirring by keeping the Magnetic Stirrer away from the pump. The reason for this can be found here.

    The magnetic stir bars mentioned for use within the syringe are actually stir discs! The thin, Parylene-coated Neodymium discs work well at a distance and do not interfere with the dispensing of fluid from the syringe. They are autoclavable, and priced so that they can be disposable. See our webpage for more information on these and other magnetic stir discs from V&P Scientific.

    This combination of V&P's innovative magnetic Tumble Stirrer and unique magnetic Stir Discs assisted Evan Macosko and the McCarroll lab at Harvard develop a method that, to paraphrase the title of their paper, profiles the genome-wide expression of individual cells on a large scale. We would love to help you do something similar with your research!

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  2. Stirring Cells or Particles in 50ml Tubes with New Stir Elements

    V&P Scientific has recently added a new line of stir elements which are capable of re-suspending cells or particles and keeping cells or particles in suspension without causing cell damage.

     The VP 772FN-25-55TF-150 {Tuning Fork} and the VP 772FN-20-21CV-150 {Torpedo-Cross} were both designed to be autoclavable. They fit inside 50ml centrifuge tubes to mix the constituents. The outer shape of both stir elements matches the inside topography of the tube.  Both designs allow you to take samples while the tube is being stirred. The Torpedo-Cross design has a lower profile and therefore requires less dead-volume than the tuning fork design. Both stir elements use two VP 782N-12-150  Neodymium Iron Boron magnetic discs.

    These stir elements have optimal performance with our VP 710D2 Multi Stirrus™. Both designs are well suited for mixing different solutions, resuspension of cells, or dissolution of powders. When these stir elements are used in conjunction with the VP 710D2 Multi Stirrus™ more than 24 50ml conical tubes can be mixed simultaneously. The stirring action does not shear or damage fragile cells even after continuous mixing.

     In a test experiment using human promyelocytic leukemia cells, 35ml samples were removed from a tissue culture flask and placed into three different 50ml conical tubes.

    The 1st tube was allowed to sit at room temperature with no stirring. The 2nd tube was stirred at 350 RPM with the Tuning Fork design{VP 772FN-25-55TF-150}. The 3rd tube was stirred at 350 RPM with the new Torpedo-Cross design {VP 772FN-20-21CV-150}.

    The cells in each tube were counted within 5 minutes of being removed from the tissue culture flask. They all had comparable cell counts with percent viability greater than 98%, as measured by Trypan blue exclusion. The three tubes were then allowed to incubate at room temperature for approximately one hour, with 2 tubes being continuously stirred at 350 RPM during the incubation.

     The cell counts taken from the tubes being stirred remained consistent with the counts that were taken within the first 5 minutes of removal from the tissue culture flask. The counts from the tube with no stir element showed a drop of 40% due to the settling of the cells. 

     The three tubes were then centrifuged for 5 minutes at 500 RPM to pellet the cells. The 2 tubes with stir elements were returned to the stir device and stirred for 5 minutes in an attempt to resuspend the pelleted cells. The counts from the tube with no stir device dropped by over 99%, consistent with the pelleting of the cells. The tubes being stirred returned to the counts comparable to those taken within the first 5 minutes of removal from the tissue culture flask. There was no evidence of cell damage seen in any tube as measured by Trypan blue dye exclusion; the percent viability remained greater than 98% in all three tubes.

     In a second test experiment, both stir elements were used to mix heavy glitter particulates. The Torpedo-Cross style element was superior in keeping heavy particulates in suspension. When very light particulates {fine glitter} were tested, the Tuning Fork design was equally effective. The Torpedo-Cross stir element, being a two piece design, is more expensive than the single piece Tuning Fork stir element, but offers a greater performance when stirring heavy particulates.

     In summation, both the Torpedo-Cross and the Tuning Fork style stir elements provide a safe and effective way of keeping cells or particulates in a uniform suspension throughout a 50ml conical tube, all the while allowing you to take samples while the tube is being stirred. The Torpedo-Cross is superior when using heavy particulates or clumping cells.

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  3. Mixing E-Liquid or E-Liquid for Vaping

    There has been an increasing amount of customers purchasing our stirrers and stir bars for mixing E-Juice. E-Juice consists of a few simple ingredients which include, Vegetable Glycerine, Propylene Glycol, Nicotine and Flavoring. Vegetable glycerin, which serves as the base of E-Liquid, has a viscosity of 1412cp. For perspective, water has a viscosity of 1cp, while honey has viscosity of 10,000cp. Magnetic mixing is an effective and efficient way to mix E-Liquid, which results in faster steeping times and less work.

     

    Standard stir plates are not magnetically strong enough to mix such a viscous liquid in larger quantities. There are many YouTube videos demonstrating magnetic mixing with homemade stirrers and/or standard stir plates. However, these smaller stir plates only work for small volumes. Our customers are not making E-Liquid for themselves, rather are producing large quantities and batches to sell.

     

    V&P Scientific collaborated with San Diego based Vape Corner. Vape Corner is locally owned and has two locations in San Diego. As their business has grown, owner Gino Sugapong has researched other methods of mixing his E-Liquid. Currently they have been mixing their E-Liquid by hand. This yields a rather physical task and a 4 week steeping time.

     

    Because the ingredients in the E-Liquid have different viscosities, they separate from each other during the initial week of “steeping”. It’s important that each batch is mixed every day so that the entire mixture is homogenous. We aimed to compare stirring magnetically vs stirring manually - shaking by hand .

    We mixed 3 – one gallon batches of the same E-Liquid. Each bottle contains: Propylene glycol, Vegetable glycerine, flavoring, nicotine

    Bottle 1.            Control: Shaking by Hand, once a day for a week

    Bottle 2.            Magnetic mixing for 30 min, once a day for a week - 210 minutes of mixing total in a week

    Bottle 3.            Magnetic mixing 24hrs every day for a week - 10,080 minutes in a week

    We used the following V&P Products:

    VP 710C5 - Stir Base

    VP 772M-N52-27-111-PTFE - Stir Paddle

       

    We know that it takes up to 4 weeks for the E-Liquid to steep if shaken by hand. Our expectations were that Sample 3 - magnetic mixing 24/7-  would yield the best results in regards to taste and decreased steeping time.


    Day 1

    Bottle 1: Hand mixed for 30 min

    Bottle 2: Mixed magnetically with paddle for 30 min.

    Bottle 3: Mixed magnetically with paddle 24 hrs.

    Notes: After 15 min. of mixing, Bottles 2 and 3,  fully emulsified. Visually, a milky color due to the introduction of air during mixing. See Photo below:

    From left to right, Bottle 1, 2, 3 after 30min


    Day 3

     

     

    Bottle 1: Bottle is slightly lighter than the rest.

    Bottle 2: Looks similar to bottle 3

    Bottle 3: Still being mixed, looks similar to bottle 2

    Notes: It appears that bottles 2 & 3 are slightly darker in color than bottle 1.

     

     See Photo Below:

    From left to right, Bottle 1, 2, 3 after 72hrs

     


    Day 4-7

    Nothing changed visually from day 3. We could arguably see there was a slight difference in color. Both bottle 2 and 3, the magnetically stirred bottles were darker. The difference was slight and could be light reflections. At day 7, we took samples from each bottle and issued a blind taste test to the employees at Vape Corner.

     

    Results:

    Bottle 1 - Hand Shake- : Flavor has not yet steeped. Still needs more time to steep.

    Bottle 2 - Magnetically Stirred for 30min/day for 7 days- : Steeping time was indistinguishable to a properly steeped batch. Flavor was  optimal.

    Bottle 3 - Magnetically Stirred 24/7 for 7 days- : Flavor was less than that of the other two bottles. However, the noticeable difference was that the Vape of this bottle was smoother the other two.

     


    Summary:

    Surprisingly enough, bottle 2 was the best outcome. We were able to achieve an appropriate steeping in one week by mixing the E-Liquid 30min/day for 7days. The benefit of this is that you won’t have to plan so far in advance for your stock of E-Liquid. This is especially true of stores, like Vape Corner, that carry a large variety of flavors. You can stock a fixed amount of any flavor, and if that begins to sell out quickly, you can make more in 1 week. This is a better scenario than having to plan 4 weeks in advance and having to tell customers you’re out of stock of their favorite flavor.

    Consequently, bottle 3 - magnetically stirred 24/7-  was smoother to inhale, though the flavor was decreasing. It is interesting to find that the increased stirring would diminish the flavor. If a smoother vape is preferable, it’s quite easy to adjust the recipe with more flavoring to compensate for the effect of the magnetic stirrer.

     

    Alternatively perhaps we should have magnetically mixed for 210 minutes to achieve the optimal flavor and done the taste test at this time point.  This would have significantly reduced the time to make a new batch.

    Using the VP 710C5 and the VP 772M-N52-27-111-PTFE, you can mix multiple bottles at the same time. Using our Patented Stirring Technology, you can quickly and easily mix viscous solutions such as E-Liquid. We were able to subjectively identify improved taste and quality of the E-Liquid with magnetic mixing versus mixing by hand.  

    Click here for videos of magnetic stirrers mixing e-liquid.

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  4. Mixing in Small HPLC Vials

    V&P Scientific makes miniature magnetic stir discs which are encapsulated in PVDF or PEEK. These magnetic discs are well suited for stirring inside small HPLC vials.

    Thomson SINGLE StEP™ and eXtractor 3D™ filter Vials are designed to speed up sample prep and analysis. The plunger filter with different membranes nestles into the vial while simultaneously filtering and readying the sample for any autosampler. This is a process that minimizes any loss of sample eliminating multiple transfers. Thomson filter vials are compatible with most standard autosamplers; such as Agilent® and Waters® including the UHPLC™.

    This photo shows a Thomson eXtractor 3D™ filter vial next to a couple of V&P PVDF encapsulated magnetic stir discs.

    These stir discs have optimal liquid mixing performance with our VP 710D2 Multi Stirrus™.  This design is well suited for mixing different solutions or dissolution of powders.

    When used in conjunction with the VP 710D2 Multi Stirrus™, more than 100 HPLC vials can be mixed simultaneously.  If resolubilizing your compounds after refrigeration, or aiding in the initial solubilization of compounds is important for your procedure, then this is an easy solution for you.

     This video demonstrates stirring in both chambers, and stirring in a single chamber after the filtration step has been completed.

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