Bacteria-free medicine: How horseshoe crabs’ blue blood saves lives

Amanda Edward
6 July 2014

Above: Horseshoe crab in sand (Wikimedia Commons/US Fish and Wildlife Service/Perry Bill)

During your lifetime, you’ll probably receive a number of vaccines to protect you from diseases such as measles, polio, and Hepatitis B. But how can you be sure that these vaccines, which are injected directly into your bloodstream, are free of bacterial contamination? After all, vaccines are designed to protect, not harm you.

Fast fact: Horseshoe crabs are used as a source of coagulogen for the pharmaceutical industry and as bait in commercial whelk (conch) and eel fisheries.The next time you get a shot, you can thank a primitive and prehistoric organism called the Atlantic horseshoe crab for your peace of mind. Actually, the name “crab” is misleading, since they are more closely related to arachnids (such as spiders) than they are to crabs and other crustaceans. But what’s most important is that their light blue blood can help keep your own blood safe!

When you get a cut, your white blood cells spring into action and try to kill as many invading pathogens (disease-causing organisms) and other foreign bodies as they can. But when a horseshoe crab is injured, its immune system dispatches amebocytes. These small, mobile cells release a substance called coagulogen to fend off invaders by coagulating (clumping together) bacteria into a gel-like substance. This immobilizes the invading bacteria and prevents them from spreading to other parts of the horseshoe crab’s body.

Coagulogen is so sensitive that it can detect bacteria at a concentration of one part per trillion. That’s like trying to find one specific coin in a pile of a million million (1,000,000,000,000,000) coins! This impressive defense mechanism has helped the horseshoe crab to survive as a species for more than 450 million years, almost twice as long as all dinosaurs have been extinct.

Coagulogen was discovered in 1956 by American medical researcher Frederick Bang, while he was researching blood circulation in the horseshoe crab. He noticed that the animal’s blue blood coagulated when either gram-negative (bacteria with thinner cell walls) or dead bacteria were introduced into its bloodstream. The cell walls of gram-negative bacteria contain endotoxins, which are only released when the bacteria die.

Fast fact: Fibrinogen is the clotting factor responsible for healing cuts and wounds in mammals, the same role played by coagulogen in the Atlantic horseshoe crab.Endotoxins are essentially chains of fat and sugar molecules that trigger an immune response. This response can range from fever to septic shock, which is a body-wide inflammation response to a severe infection. So you definitely don’t want to get an injection contaminated with endotoxins.

But endotoxins are also very heat stable, which means they can survive the sterilization process used in making vaccines, intravenous medication, and medical equipment. Before coagulogen was discovered, rabbits were injected with vaccines and other medication to test for the presence of these harmful bacteria, a process that was very time-consuming and not very much fun for the bunnies.

Thanks to the discovery of coagulogen, scientists were able to develop the Limulus Amebocyte Lysate (LAL) test in the 1970s. This quick, efficient, and fool-proof test ensures that intravenous medicines and related equipment are sterile (completely free of harmful bacteria). The blood of the horseshoe crab is required for this test, but after they make their donation the animals are returned to the wild.

The horseshoe crab blood is then centrifuged (spun around quickly) to separate the amebocytes from the rest of the fluid. Next, the amebocytes are lysed (burst open) to release the coagulogen, which is used to detect endotoxins.

Fast fact: Human blood is red because of a protein called hemoglobin, which contains iron atoms. Atlantic horseshoe crab blood is blue because of hemocyanin, which contains copper atoms.If you were born after 1970 and you live in North America, every single vaccine or intravenous fluid you have ever been given, and any piece of medical equipment that has come in contact with your blood would have passed a one-hour LAL test. A simple test tube inversion (turning the tube upside down) confirms if a gel has formed, indicating bacterial contamination. If the sample is sterile, it remains a liquid.

Although the blood donation does not appear to have an immediate effect on most horseshoe crabs, it appears that between 10% and 30% die shortly afterward. Researchers know this because they tag and track the animals. Additionally, fewer and fewer horseshoe crabs are returning to their spawning grounds each year, and those that do return move more slowly and have less protein in their blood (their blue blood is getting thinner). In fact, a research team studying the mortality of Malaysian horseshoe crabs in a laboratory setting discovered that 17.5% died within 60 days of being bled.

Fortunately for the horseshoe crabs, researchers are developing other tests that could replace the LAL test. These include synthesizing (creating) a liquid crystal that also clumps in the presence of endotoxin, as well as using an electronic chip that emits a signal when bacteria are detected! But until one of these tests is shown to be effective and reliable, manufacturers of vaccines, medicines, and medical equipment will need to continue collecting the light blue blood of this ancient animal.


General information

The Annual Blood Harvest of the Horseshoe Crab (Wired UK/Olivia Solon) The Blood Harvest (The Atlantic/Alexis C. Madrigal) Blue Blood Donors of the Sea (The Sterile Eye/Øystein Horgmo) Medical Uses: How does the horseshoe crab protect the public health? (The Horseshoe Crab/Ecological Research & Development Group) New sensor derived from frogs may help fight bacteria and save wildlife (EurekAlert!/Chris Emery)

Scholarly publications

Akbar JB, Jalal CA, Zaleha K, Armstrong P, Kmaruzzaman BY. 2011. Effects of blood extraction on the mortality of Malaysian horseshoe crabs (Tachypleus gigas). Marine Behaviour and Physiology. 44, 5: 321-327. Anderson RL, Watson WH, Chabot CC. 2013. Sublethal Behaviour and Physiological Effects of the Biomedical Bleeding Process on the American Horseshoe Crab, Limulus polyphemus. Biology Bulletin. 225: 137-151. Odell J, Mather ME, Muth RM. 2005. A Biosocial Approach for Analyzing Environmental Conflicts: A Case Study of Horseshoe Crab Allocation. BioScience. 55, 9: 735-748. Osaki T, Kawabata H. 2004. Structure and function of coagulogen, a clottable protein in horseshoe crabs. Cellular and Molecular Life Sciences. 61: 1257-1265. Rudloe A. 1983. The effect of heavy bleeding on the mortality of the horseshoe crab, Limulus polyphemus, in the natural environment. Journal of Invertebrate Pathology. 42, 2: 167-176.

Amanda Edward

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