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CELL CULTURE'S SPIDER SILK ROAD
 
Jeffrey Perkel
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Golden orb spiders are large—about the size of the palm of the hand—but they rarely bite and are relatively harmless to humans. To collect their silk, Kuhbier would nudge a spider from its web into a plastic box, immobilize it on a Styrofoam cube (without harming it), and gently pull the dangling dragline fiber from the animal's abdomen, winding it around a spindle. Kuhbier says he could pull up to 200 meters per animal in about 45 minutes, at which point the spider was returned to its web. A YouTube video on the Malagasy textile illustrates how delicate that process can be: “You can just see these diaphanous spider silk strands kind of floating around the hands of these individuals,” a museum curator explains. [2,]

At first, Kuhbier did his silking manually; later, a motorized device was built in collaboration with the University of Hannover that extracted and wound the fiber more regularly and evenly—an idea that, as the team discovered after two years of work, actually dates back to the eighteenth century, when a Spanish monk in Madagascar described perhaps the first ever spider silk-harvesting apparatus.

Collecting the silk was not the only hurdle though. “Spider silk has the ability to supercontract when it comes in contact with water.” In aqueous solution—like, say, the culture media cells are grown in—the material knots up into an unusable bundle. Kuhbier needed a way to keep the silk taut.

It took him about a year and a half to develop a solution. Dentists, he recalled, often use an inexpensive bendable wire to hold dental prosthetics in place. Why not use that wire to create a “weaving frame” for spider silk? Such an assembly should prevent the silk from clumping, thereby retaining its structure when submerged in aqueous solutions.

The end result was a tiny golden silk tapestry 5 or 20 mm on a side—a stamp-sized golden miniature of the textile in New York. The structures are highly stable, he says—they can be washed in ethanol and autoclaved, and also seem to be resistant to both bacterial and fungal degradation. And they are small enough to fit in the wells of 6-well microtiter culture dishes. When Kuhbier seeded those frames with NIH 3T3 fibroblasts, he found that the cells not only adhered to the fibers (which are visible in photomicrographs due to their strong autofluorescence) but also proliferated, migrated, and thrived. [3,] “They were in a healthy state and vital.”

The frames also support true 3-D cell culture. In 2011, Kuhbier co-authored a study in which his weaving frames were used to build “artificial skin.” [4,] Here, the frame was stuffed with a ball (or “clew”) of silk fibers to create a contiguous 3-D structure sandwiched between the two sides of the weaving frame. The silk frame was populated first with mouse embryonic fibroblasts and later with a surface layer of keratinocytes, which were grown at the liquid-air interface to force a more epidermal configuration. The resulting structure, he says, “formed skin-typical markers [visible] in immunofluorescence. So it seems to be a very good matrix for cell culturing.”





Wrangling Silk Genes

But that doesn't mean the silk is convenient to use. Maintaining and silking live spiders is laborious and— Malagasy silk weavers notwithstanding—unscalable. As a result, many researchers concentrate on the silk genes themselves, mixing and matching protein segments to produce materials with the properties they desire.

As a graduate student, Anna Rising, now Associate Professor of Neurobiology at the Swedish University of Agriculture Sciences and the Karolinska Institute in Stockholm, decided to clone the gene encoding dragline silk protein from the nursery-web spider, Euprosthenops australis. Dragline silk forms the outer edges and structural support of a spider's web; it is the silk spiders use to catch themselves when they fall. E. australis produces an exceptionally strong dragline, with a tensile strength of 1.5 GPa. But almost nothing was actually known about the organism itself, including where to find it, how to catch it, and whether or not the creature was poisonous.

Nevertheless, Rising traveled to South Africa to bag herself some spiders. Here's the thing, though: It's not that easy to find a spider in the wilderness. She and her guides first tried using headlamps to reflect the spiders’ eyes. But there are many animals in the bush, and many sets of eyes. Eventually, they realized their best bet was to search for the animals’ distinctive funnel-shaped webs, which extend from a tree to the ground, where the spider waits in a burrow.

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