The overall goal of this procedure is to create an inexpensive and effective filter for concentrating a wide variety of waterborne pathogens from large volumes of water. Filter construction is achieved by first rinsing glass wool, washing it with both an acid and a base, and re rinsing the wool to a neutral pH. Next, the glass wool is rinsed in phosphate buffered saline.
In the next step, the glass wool is packed into a PVC pipe to form the glass wool filter. After using the filter for sampling water, an elution step is performed. The sample can then be stored, concentrated further, or proceed to be analyzed for pathogens by cultural or molecular methods.
Waterborne disease is a significant threat to public health. Worldwide disease transmission can result from drinking water and recreational water activities. One approach for evaluating the sanitary quality of water is to test it directly for disease causing pathogens.
There are several benefits for using glass wool filters as opposed to other traditional waterborne pathogen filters. First, the filters are inexpensive, allowing for a large number of samples. Second glass wool allows the concentration of a large volume of water, which is often necessary for detecting waterborne pathogens.
Third glass wool filters are also very portable, making it easy to conduct field sampling. In remote locations, glass wool filters are capable of concentrating a variety of pathogens, including viruses, bacteria, and protozoa. Finally used in conjunction with a pre-filter.
Glass wool filters will concentrate pathogens in difficult water matrices. The materials required for making glass wool filters Are safety equipment, gloves, eyewear, and a protective gown. Glass wool, a sterile autoclave bucket, one molar hydrochloric acid, and one molar sodium hydroxide pH strips, or a pH meter, a timer Teflon taped threaded PVC pipe.
In this demonstration, we use two inch by four inch, two inch screw on caps with Teflon taped barbed couplings, polypropylene mesh perfil, a 60 milliliter syringe, metal plungers sterile containers, a scale. You will also need sterile reverse osmosis water and sterile phosphate buffered saline before and after making each batch of filters. Sterilize your work area with 10%bleach solution.
To begin this procedure, place the glass wool in a sterile autoclave bucket. Wet the glass wool with sterile reverse osmosis water when completely saturated, allowed to sit for 15 minutes. Drain the water from the bucket.
Saturate the glass wool with one molar hydrochloric acid mix, and let sit for 15 minutes. Drain the hydrochloric acid. Rinse the glass wool with sterile, reverse osmosis water.
Mix thoroughly and check the pH. The rinse may need to be done several times to attain a neutral pH. Pour off the rinse water.
Saturate the glass wool with one molar sodium hydroxide mix, and let sit for 15 minutes. Pour off the sodium hydroxide. Repeat the sterile reverse osmosis water rinse again.
The rinse may need to be repeated several times until the pH is seven. Drain the rinse water. Cover the glass wool completely with sterile phosphate buffered saline.
This can be stored at four degrees Celsius, but check the pH before use to make sure it is neutral. The pH of the solution will rise over time and might need to be re rinsed with phosphate buffered saline. In this video, we have shown the glass wool preparation in a bucket, but if many filters are required, faster washing systems can be used, such as this one which has a convenient bottom valve for easier draining and pumping of water.
For rinsing. In this demonstration, we use two inch by four inch threaded PVC pipe and two inch screw-on end caps with half inch barbed couplings. Other sizes may be used.
Apply Teflon tape to threaded portions for a watertight seal. Take small pieces of glass wool and begin packing the filter. Make sure the glass wool fits tight around the edges and try to avoid channeling packed tight with a metal plunger for the pipe size shown here.
85 grams washed and packed glass wool is used per pipe. As an alternative, a hand press may be used. Insert polypropylene mesh into the ends of the filter.
Screw on the threaded caps. Make sure that connections are tight to avoid leaking. Label the sample inflow and outflow ends of the filter.
Push 60 milliliters of phosphate buffered saline into the glass wool filter. Using a catheter tipped syringe, excess will come out the opposite end, wrap the ends tightly with parfum to keep the filter moist. The finished filter can be stored for up to 30 Days at four degrees Celsius.
When sampling, If the water pH is greater than 7.5, it must be adjusted to between 6.5 and seven with hydrochloric acid. Otherwise, virus absorption to the glass wolf fibers will be poor, and a false negative sample might result in this demonstration. A precision peristaltic pump is used to add acid to the sample inflow line.
Instead of a pump, you could use an inexpensive Venturi tube depending on the sediment level in the water, a pre-filter may be used, adjust the flow rate through the filter to between two and four liters per minute. Typical sample volumes for this method are between 200 to 1500 liters. Disconnect the filter and place in a sterile plastic bag.
Keep the filter at four degrees Celsius for up to 48 hours After a sample has been Taken, affix the filter to a ring stand with the barbed inflow coupling pointing downward into a polypropylene bottle. Push 80 milliliters of 3%beef extract with 0.05 molar glycine with a pH of 9.5 into the filter, allowed to sit for 15 minutes. Push another 80 milliliters of beef extract through the filter, followed by air until foam comes out of the inlet.
Adjust the pH to between seven and 7.5 with one molar hydrochloric acid. This can then be stored at four degrees Celsius for up to 24 hours or negative 20 degrees Celsius for longer periods of time. In this video, you have learned how to construct a glass wool filter for concentrating a variety of pathogens from large volumes of water.
Glass wool filters have been shown to be effective in concentrating human viruses such as enterovirus, adenovirus, and norovirus, as well as a variety of bovine and teric viruses and avian influenza virus. The filter will also concentrate bacteria such as e coli, salmonella, and campylobacter species, and proteasome parasites such as Cryptosporidium and Giardia.
