Hoefer SG500
Contents
1. Note The entire solution remaining in the mixing cham ber is at the final concentration and does not contribute further to the gradient If desired it can be used as a displacement solution for convex gradients unless it contains acrylamide Q Close the slide valve and add a portion of the final solution to the reservoir back chamber Carefully open the slide valve and allow just enough solution to flow through the connector channel to fill it to the edge of the mixing chamber then close the valve Be sure no large bubbles remain to obstruct flow through the channel Add the required volume of the starting solution to the mixing chamber With the stopper tubing open insert the stopper assembly securely into the mixing cham ber Fig 4 Clamp off the tubing on the vent airtight O Start the magnetic stirrer and unclamp the outlet tubing if needed Simultaneously open the slide valve and start the pump Stop the pump and remove the tubing from the receiving container as soon as the last of the solution has left the reservoir chamber Disconnect tubing Flush and rinse all parts thoroughly with distilled water after use 3 Care and maintenance The gradient maker should be cleaned thoroughly with distilled water after use to prevent polymerization or crystallization of solutions in the chambers and stopcocks Do not use abrasives acetone pure alcohols or organ2. volume res reservoir chamber mix mixing chamber Fig 3 Examples of exponential gradients relative concentration 0 20 40 60 80 100 of gradient delivered To generate an exponential gradient 0 Make sure all parts are clean and liquid flows freely through all channels stopcocks and tubing 2 Assemble the stopper Select a one hole rubber stop per that fits securely in the top of the mixing cham ber Attach a short piece of flexible vinyl tubing that can be clamped off to make an airtight seal to a rigid piece of glass or plastic tubing that fits tightly in the hole of the stopper The stopper assembly should seal the mixing chamber airtight when the slide valve is closed and the outlet tubing is clamped off Add a magnetic stir bar of the appropriate dimen sions 20 30 mm long to the mixing chamber and place the unit on a magnetic stirrer Connect tubing to the outlet connector and pump then adjust pump speed if needed Position or connect the tubing to the receiving vessel gel casting unit centrifuge tube etc at top or bottom as appropriate Fig 4 Gradient maker with plunger in place Note If the gradient volume is larger than the reservoir will hold the additional volume can be added during the gradi ent delivery Do not allow the reservoir chamber to go dry as bubbles will be pulled into the mixing chamber changing the dilution volume and gradient shape
3. OD rana Hoefer SG500 Gradient maker um SG500 IM Rev E0 08 12 Ayo e fe r Page finder SG500 instructions 1 nennen 1 1 Generating linear gradients unne 2 Pouring gradients from the top nnn enn 4 Pouring gradients from the bottom 5 2 Generating convex and concave exponential PNAC SMES sich een anne ae nee AEE 6 To generate an exponential gradient 7 3 Care and maintenaNC ssi airesin 9 A Ordering information uenseesssseneseeneneenenennenennnn 9 Fig 1 Hoefer SG500 gradient maker Included but not shown Hose clamps 4 SG500 instructions The Hoefer SG500 gradient maker is designed for producing linear gradients of aqueous solutions ranging in volume from 100 500 ml The SG500 can be used to create convex and concave exponential gradients with the addition of a one holed rubber stopper a piece of rigid tubing and a piece of flexible tubing The unit is fabricated from cast acrylic plastic sheet and tubing It is suitable for casting acrylamide pore gradient gels and delivering salt gradients for low pressure chromatography systems reservoir back chamber mixing front chamber outlet connector 4 mm ID tubing slide stop push pull valve connector channel open closed Fig 2 Open and closed positions of slide valve 1 Generating linear gradients To generate a linear gradient between two concentrations equal vol
4. e If there is a substan tial difference in densities between the two solutions there will be a sudden flow from the denser chamber to the lighter chamber to bring the two into hydrostatic balance This will result in the gradient not being fully linear To avoid this add equal weights rather than volumes of the solutions to the appropriate chambers Q Carefully open the slide valve and allow just enough solution to flow through the connector channel to fill it to the edge of the mixing chamber then close the valve Be sure no large bubbles remain to obstruct flow through the channel Add the required volume of the starting solution to the mixing chamber and start the magnetic stirrer Open the outlet tubing if clamped off o Simultaneously start the pump 8 If it is important that no bubbles disturb the gradient watch the delivery carefully and as soon as the last of the solution has entered the pump head stop the pump and clamp off the tubing to the receiving container Flush and rinse all parts thoroughly with distilled water after use Pouring gradients from the top Filling a container with a gradient e g casting acrylamide pore gradient gels can be done either dense solution first from the top or light solution first from the bottom To fill from the top Proceed as described in Section 1 putting light final or top solution in the reservoi
5. e gradient mix has been delivered It is convenient to include a dye in the displacement solution to visually track the boundary between the gradient mix and the displacement solution 2 Generating convex and concave exponential gradients By holding the volume in the mixing chamber constant as opposed to declining for linear gradients the gradient generated will have an exponential concentration curve The mixing volume in the SG500 unit can be held constant by sealing the top of the mixing chamber with a rubber stopper and tubing assembly Fig 4 The total volume of the gradient can be greater than the maximum volume of the gradient former V because additional solution can be repeatedl Ce Vres Cri Va Va P y Ves KORTEN examples of the gradients generated by various res added to the reservoir chamber Using Eq 1 combinations of volumes and concentrations listed in Table 1 are plotted in Fig 3 Concave gradients are generated high concentration first and thus containers such as gel cassettes must be filled from the top Convex gradients must be filled from the bottom Most frequently used is the concave pore gradient for increased reso lution in the high molecular weight region of acrylamide gels Table 1 Examples of concave and convex exponential gradients A B c D E Cres 0 1 1 1 0 5 Vres 500 500 100 500 500 Cmix 1 0 0 0 5 1 Vmix 100 100 500 100 100 Volumes are relative C concentration V
6. ic solvents to clean this unit 4 Ordering information product qty code no SG500 Gradient Maker 1 SG500 500 ml total volume Push pull valve for salt gradient 1 SG500 10 Red outlet fitting 4 mm 4 XPO10 Push pull valve replacement 1 SG500 1 Hoefer Inc 84 October Hill Road Holliston MA 01746 Toll Free 1 800 227 4750 Phone 1 508 893 8999 Fax 1 508 893 0176 E mail support hoeferinc com Web www hoeferinc com Hoefer is a registered trademark of Hoefer Inc 2012 Hoefer Inc All rights reserved Printed in the USA Hoefer
7. r chamber step 3 and dense solution in the mixing chamber step 5 2 Place the delivery outlet against the upper edge of the receiving container Adjust the pump rate so that the solution flows evenly down the side in a smooth continuous stream The delivery speed should be slow enough that the newly arriving solution does not mix with the underlying solution Alternatively using a rigid cannula at the end of the delivery tubing hold the tip of the cannula just above the surface of the solution raising it smoothly as the container fills Pouring gradients from the bottom This technique is commonly used in filling multiple gel casting chambers 0 Proceed as described in Section 1 putting dense final or bottom solution in the reservoir chamber step 3 and light solution in the mixing chamber step 5 2 In step 2 connect the outlet tubing to the bottom inlet of a gel casting unit or to a cannula long enough to reach the bottom of the receiving container Adjust the pump rate so that the solution is not forced up in a fountain that mixes with the overlying solution If all of the gradient solution must be delivered to the container a displacement solution may be used Just as the last of the gradient mix is pumped out of the mixing chamber and before any air enters the tubing add an appropriate volume of a denser displacement solution to the mixing chamber and pump it through until all of th
8. umes of solutions of the two concentrations are measured into the two chambers of the gradient maker As solution is delivered out of the mixing chamber an equal volume flows in from the reservoir chamber where it is rapidly diluted and mixed to unifor mity by a magnetic stir bar The initial concen tration delivered will be that of the solution in the mixing chamber the final concentration will be that of the reservoir chamber For the most consistent delivery of gradients a peristaltic pump is recommended Make sure all parts are clean and liquid flows freely through all the connecting channel of the slide valve and pump and delivery tubing 2 Add a magnetic stir bar of the appropriate dimensions 20 30 mm long to the mixing chamber and place the unit on a magnetic stirrer If volumes will be less than half the capacity of the unit an identical stir bar should be placed in the reservoir chamber as well to balance the displacement and prevent backflow into the reservoir when the chambers are first connected Connect tubing to the outlet connector and pump and adjust pump speed if used Position or connect the tubing to the receiving vessel gel casting unit column etc Close the slide valve out on side of white slide stop button Fig 2 If the outlet tubing is not controlled by a pump clamp it off near the gradient former Add the required volume of the final solution to the reservoir back chamber Not
Download Pdf Manuals
Related Search