Can any one please give me idea how we can load less sample in a loop? For an example we have a 10mL loop but we have 5 mL of Sample but we do not want to dilute the sample with WFI or any other Buffer, how we can inject the sample without any bubble in the Column in the AKTA Purifier/Explorer.
In your case, the only solution is to replace your 10 ml loop with 5 ml loop.
The manufacturer sells injection loops in different volumes for this exact application (0.500 ml, 1 ml, 2ml, 5ml, 10ml, 20 ml...). Loops are meant to be switched out to best suite your own application and it is a good idea to have several sizes on hand. Purchase a few different sized loops so you will have more injection options in the future (they are inexpensive and you can make your own loops if you have some tubing, proper fittings and an analytical balance to calibrate them).
Purchase in here in fact time consuming. Actually, right now I need to process 5 mL sample but I have 10 mL loop rather 5 mL. However Thanks for your suggestion.
William, could you please tell me how can I make my own 5mL loop with proper calibration as I have loop of 75 mm and 1 mm diameter. It would be appreciated to have your potential answer.
Easy. Cut your 10ml loop in half. Check the volume by measuring how much water it holds. *Seriously, I really do not recommend cutting it in half, unless you have no choice.
Better to make your own injection loop(s). Use high quality HPLC grade SS (or PEEK if applicable) tubing only. The size of the tubing is not critical, but it should be much longer than wide due to diffusion and mixing concerns. In your case, you need to create a length of tubing with a volume of 5.000 mls inside. Water has a density of 1 gram per ml so use water to calibrate a cut piece of tubing by weighing the amount of water it contains. This will give you a volume within the accuracy of your balance.
Start by estimating the length of tubing needed to hold 5 mls by calculating the volume found in the tubing you have.
Volume of a cylinder = π r2 h
1mm ID tubing contains about 8.103 ul of liquid per cm of length. Your 75mm long tube would hold about 60 ul of liquid so you can not use it to make a 5 ml loop. 5,000 ul would use 617 cms of 1 mm ID tubing! IOW: Use wider ID tubing for this. A 5mm ID x 25 cm length of tubing will hold just over 4.9 ml. Because tubing ID is not uniform, cut a length longer than needed, fill it with water, weigh it and cut off small sections to get down to the size you need. Carefully cut/file the ends perfectly square and smooth (under magnification), add proper fittings and install.
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