Horseshoe Crabs and their Surprising Role in Vaccine Development
If you’ve ever been to a beach on the east coast, vacationed in Cancún one summer, or simply visited the Houston Museum of Natural Science, you may have seen a uniquely pre-historic-looking critter that still thrives today! The horseshoe crab is in fact not a proper crab—not even a crustacean for that matter. With its unique three-segmented structure, it has barely changed over the last 200 million years, surviving countless devastating extinction events such as the end-Permian mass extinction, which led to the extinction of more than 81% of marine species.
So how did these “living fossils” survive for so long? Firstly, horseshoe crabs can eat almost any organic matter, ranging from clams to algae to decaying fish. Secondly, horseshoe crabs can survive without food for up to a year. Thirdly, horseshoe crabs can survive in a wide range of salt and oxygen levels, allowing them to adapt to changing ocean conditions over the years.
However, this last point in question may start to change due to a rather surprising use of horseshoe crabs by humans—in vaccine development! It all started in 1885, when scientists at Johns Hopkins University discovered that local horseshoe crabs off the Maryland coast, the Atlantic horseshoe crab or Limulus polyphemus, noticed that the crabs’ blood would coagulate—or harden—though they were unsure about what exactly caused this hardening. Eventually, researchers would discover that the Limulus blood would coagulate in the presence of bacterial endotoxins, substances produced by gram-negative bacteria that happen to be lethal to humans.
Suddenly, new possibilities arose from the discovery. You see, drugs and vaccines can often be contaminated with bacteria, which can pose risk to humans when they release endotoxins. Thus, in order to test these drugs and vaccines, scientists had to use a variety of time-consuming and expensive pyrogen tests, such as the rabbit pyrogen test in which rabbits would be injected with samples of the endotoxin and observed for fever responses. Unfortunately, the rabbits would pretty much always die after exposure to the endotoxin, which meant that scientists had to keep replacing rabbits when testing new vaccines or drugs for endotoxins. The new Limulus amebocyte lysate (LAL) extraction process, on the other hand, only resulted in the death of up to 30% of horseshoe crabs. Additionally, the new test resulted in considerably less false negatives and more true positives than the rabbit pyrogen test.
Until 2025, LAL extracted from horseshoe crabs remained the preferred method of endotoxin testing by the United States Pharmacopeia, in charge of setting standards of pharmacological tests in the US. However, the alarming decrease in horseshoe crab numbers have been noticed since the widespread adaptation and growth of the LAL extraction method, including in Asia with horseshoe crab blood extracted from other species of horseshoe crab, called the Tachypleus amebocyte lysate (TAL) test. Scientists have since adopted and recognized the recombinant factor C (rFC) assay, a synthetic substrate that lights up when endotoxin binds to it, which has begun to replace horseshoe crab blood in the pharmaceutical industry—thereby both preserving the millions of years-old species and paving a new path forward with cheaper but more accurate endotoxin tests.