DIY

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Helminthic Therapy > DIY

Contents 1 Do It Yourself Helminth Therapy 2 Egg Counts 3 Laboratory technique describing hookworm and whipworm propagation 4 Building Incubators 5 Sources for Lab Chemicals and Media 6 General Laboratory Technique 7 Materials List

Do It Yourself Helminth Therapy

This page is dedicated to documenting efforts of helminth therapy users to provide helminth therapy for themselves.

• Instructions for Incubating Hookworm Larvae - Several people share their techniques for incubating hookworms.
• Incubating Hookworm Yahoo Group - A great discussion group for DIYers.
• Counting Club - A great website about how to count eggs

Egg Counts

One of the most accurate ways to judge the degree of hookworm infection is helminth eggs in feces. The standard laboratory procedure for this is called the McMaster egg count technique.

Laboratory technique describing hookworm and whipworm propagation

• An Agar Plate Method for Culturing Hookworm Larvae: Analysis of Growth Kinetics and Infectivity Compared With Standard Coproculture Techniques. Daniel Reiss, Lisa M. Harrison, Richard Bungiro, AND Michael Cappello, American Society of Tropical Medicine and Hygiene, 77(6), 2007, pp. 1087-1090

• Incubating strongyloides Includes and explanation of the Baermann apparatus (funnel) for isolating worm larvae from media

• Canine Hookworm Incubation

see box 1, also includes description of a phosphate buffer

• Culturing Hookworm

• Incubating Hookworm Larvae A users technique

• CulturingHookworm Culturing Hookworm 2 A users a account of their technique, posted here.

• MacMaster egg counting technique

• More accounts of egg counting and incubating

• incubate_sterilize incubation and sterilization

Building Incubators

Hookworm eggs hatch best between 25-35 C (77 - 95 F) with relative humidity above 60%. Low cost incubators can be rigged from Styrofoam coolers, or if you wish to do a little more work, constructed from Styrofoam insulation from a home improvement store, and glued together with silicon sealant.

A source of humidity is also needed, which can be as simple as a pan of water in the incubator.

Aquarium stores and pet shops have several items that will be useful in constructing incubators. Resistive heaters with standard light bulb sockets are available as are infrared bulbs. Hookworms don't like light but many people report success using standard light bulbs for heaters, so this appears to be fairly unimportant. Inexpensive remote thermometers for use in fish tanks and aquariums are very useful.

Controlling the heat source is an issue. It is possible to use an "open-loop" type of control over the heat source by controlling the heat, with a light dimmer say, and simply monitoring the temperature, while very slowly raising the power level. However there is an element of danger that should be carefully considered in putting a heat source in an enclosed place and we strongly recommend one of the inexpensive thermostatic controllers made for reptile terrariums (they have a wider range than aquarium heaters and also have useful remote sensors). Here is one. Controllers with digital control and humidity reporting are available for a little more money.[| HygroTherm is one brand name. Other users have reported finding useful (and possible better-made) thermostatic controllers on ebay for about the same price as the controllers shown.

Conservative design principles would still indicate that heaters should be sized as small as possible (similar to the open-loop method), in case of thermostat failure, so that peak temperatures remain as low as possible, with the heater on all of the time.

A method of controlling humidity remotely is using a small aquarium pump and an airstone in a pan of water. The air bubbles carry humidity out of the pan much more rapidly than just evaporation. Inexpensive timers - that will control on-off cycles in as small an increment as 10 minutes are also available in aquarium or pet stores.

It may also be effective to buy an entire incubator with temperature and humidity control. This one was designed for reptile eggs but looks like it could be made to work well.

This one was designed for poultry but is used for reptiles. It is very inexpensive but would probably work fine for hookworm. It doesn't control humidity, but humidity could be made high by incubating in a semi enclosed container within the incubator.

Sources for Lab Chemicals and Media

• US Biologicals Nematode Growth Media, mentioned in the Reiss paper above[

• Mono Potassium Phosphate (KH2PO4) in one pound lots A common buffer for use in preparing petri dishes for worm culture. Used with DiPotassium Phosphate to make a buffer that will maintain slightly basic PH (eg 7.2) in growth media

• Dipotassium Phosphate (K2HPO4) Used with Mono Potassium Phosphate to make a buffer that will maintain slightly basic PH (eg 7.2) in growth media

General Laboratory Technique

• Simple pH buffers

Materials List

Suppliers and materials we have found helpful. This should not be considered a Bill of Materials for incubating hookworm, although we hope to provide a BOM eventually.

• Bone Charcoal
  

Bone charcoal is charred animal bones, which are basic in pH, unlike a lot of hardwood charcoal. For those wishing to avoid animal products, coconut charcoal, on the same site, also appears to be basic. We selected the more granular of the two grades offered, without any particular reason. Papers on laboratory technique indicate that including charcoal in the incubating mixture eliminates most smell from the process. The charcoal's pH should help the resulting culture maintain a basic pH.

• Accurate Chemical Sodium Phosphate Buffer pH 7.4

This website has a fairly minimal interface. Search for "Sodium phosphate buffer powder, 0.1M, pH 7.4"

• Inexpensive pH meter ~$30

This site also has small packets of buffer for meter calibration. The meter manufacturer tells us that, although they sell a lot these meters, it is is their low-end pH meter which only carries a 30 day warranty. There is not much of a user manual or details about care and storage. There is a special solution made for preserving the probe while in storage. Try to get this in a package deal with the instrument if you choose this meter.