"A variety of different biological structures are formed by the directed assembly of large homogeneous protein macromolecules. Many of these processes have been simulated successfully in vitro. Yet, the products of such simulations often lack the specificity and material characteristics of the structures formed within biological systems....
"Presumably, the mismatch between in vivo and in vitro products derives, in part, from biophysical differences in the assembly environments. Whereas in the in vivo assembly occurs in viscous template-directed fashion, the in vitro assembly occurs in an environment with large scale bulk fluid motions, density driven flows and shear flows. The latter assembly conditions should foster disorder during macromolecule assembly unless template constraints are very stringent." (1, p. 1)
This experiment (together with those of Stodieck, Consort 1 and related KC-135 experiments) "...focuses upon using reduced gravity environments to evaluate in vitro macromolecule assembly processes and products. The experiments were designed to evaluate the effects of the reduced gravity assembly conditions on rates of assembly and product characteristics." (1, p. 1) More specifically, this Consort 1 polymerization experiment was designed to study the low-gravity formation of collagen structures.
"Collagen...is an important protein because it is polymerized into fibers that underlie the structural properties of nearly all body tissues." (3, p. 4) During the formation of collagen in the low-gravity environment, bulk fluid convective mixing, shear forces, and material sedimentation should be reduced. Such a reduction of these gravity-induced phenomena should result in "...the slower more homogeneous formation of biomaterials." (4, viewgraphs)
The experiment was one of 17 investigations simultaneously performed in the Materials Dispersion Apparatus (MDA) on Consort 1 (see also Burgess, Consort 1; Cassanto, Consort 1; Fiske, Consort 1; Hammerstedt, Consort 1; Pellegrino, Consort 1 (three experiments); Rodriguez, Consort 1; Schoonen, Consort 1; Stodieck, Consort 1; Todd, Consort 1 (four experiments); Vera, Consort 1 (two experiments)). While most of the investigative teams (including this one) used the MDA to conduct a liquid-liquid or liquid-solid diffusion study (11 experiments), the apparatus was also used to (a) determine the role of fluid physics on the formation of films and the casting of membranes (five experiments) and (b) determine the effect of re-entry loads on terrestrial-grown crystals (one experiment).
The MDA was fashioned (in part) from two teflon blocks. Each block had over 100 sample wells strategically drilled into its surface. A well was capable of holding 100-500 ml of fluid. After all of the wells had been filled with the appropriate sample materials (depending on each investigator's specific objectives), the blocks were joined together. Twice during the mission, the upper block moved relative to the bottom block. Each movement of the block either (a) aligned or (b) misaligned wells on the top block with wells on the bottom. When the wells were aligned, material transfer from one well to the other could take place. When the wells were not aligned, material transfer from one well to the other could not take place. Appropriate positioning of the wells on the top block to those on the bottom resulted in what was called "Type 1," "Type 2," "Type 3," or "Type 4" test wells. The "Type 1" test wells were used exclusively by Cassanto et al. for a protein crystal stability experiment which investigated the effect of re-entry loads on terrestrial-grown crystals (see Cassanto et al., Consort 1). The "Type 2" and "Type 3" test wells were used for either the liquid-liquid or liquid-solid diffusion experiments. The "Type 4" test wells were used exclusively for the film formation and membrane casting experiments (see Pellegrino, Consort 1; Vera, Consort 1). This experiment was allotted two "Type 3" test wells. Discussion of this specific well-type is detailed here.
Each "Type 3" test well provided the investigator with one experimental opportunity. The well-type used three sample wells, two in the top block and one in the bottom block. Thirty-six hours prior to the rocket launch, the well in the lower block of each well-type was filled with solubilized collagen. Twelve hours later, one upper well of each well-type was filled with initiation buffer and the second upper well of each well-type was filled with 2% glutaraldehyde fixative. The long delay in the filling of the upper and lower wells "...was primarily due to the relatively large number of wells that had to be precisely loaded." (1, p. 3)
Reportedly, the fluids used in the investigation were chosen to match the opportunities/restraints of the Consort 1 rocket and the MDA. "Early" sample loading of the MDA (24-48 hours prior to launch), processing temperature of the samples (approximately 22 ¡C), high g loads experienced by the vehicle during takeoff and landing (up to 20g's), 10 -5 -10 -6 g low-gravity duration (approximately 7 minutes), and "late" retrieval of samples (approximately 4 hours post-flight) limited fluid selection and investigative objectives.
Prior to the rocket flight, the blocks were joined together such that the wells in the upper block were purposely misaligned with the wells in the lower block. This prevented materials within the wells from coming into contact prior to the initiation of the experiment. Once the rocket had been launched and the low-gravity phase had been achieved, a motor and cam mechanism moved the upper block to the right, aligning the wells containing liquid solubilized collagen and the initiation buffer. Once the wells were in contact, material diffusion was realized across the liquid-liquid interface. Just prior to the termination of the low-gravity phase, the upper block again moved right, aligning the well which originally contained only solubilized collagen (but now contained a mixture of solubilized collagen and buffer) with the well containing the liquid glutaraldehyde fixative. These wells remained in contact throughout the re-entry period of the rocket.
Post-flight evaluation of the operation of the MDA indicated that the upper block moved as expected allowing diffusion to take place in the "Type 3" test wells. Although it was reported that minor fluid leakage in the top and bottom blocks of the MDA resulted in contamination of some of the test wells allotted to the 17 investigations, it was not clear if this particular experiment was affected by the contamination.
Post-flight analysis of the contents of the wells was performed. Reportedly, "Because of steep concentration gradients across the interface of the precursor and initiator solutions, diffusion was expected to occur at relatively high rates. High rates of diffusion were essential to maximize the amount of material polymerized fibers to remain in the precursor well since fibers would exhibit very high molecular weights and thus would not diffuse away from the assembly location.
"As expected, plugs of collagen...materials...were localized to the well originally containing the precursor solutions. Plugs extended to approximately half of the 16-mm depth suggesting that this was the depth reached by diffusion initiator components. The plugs of material most importantly, were of sufficient size and quality to permit subsequent analysis steps...Since the plugs have depths that are quite large compared to fiber lengths, an important series of questions can be addressed. These questions relate to the diffusion distances supported by these periods of high quality low gravity and to any intrinsic effect that diffusion may have on the polymerization process in these sensitive reactions." (3, p. 6)
Gross observations of the opacity of the collagen indicated that significant assembly of the three-dimensional lattice had occurred (the assembly did not appear to be complete).
It was further noted that:
(1) mixing was dependent upon small convective forces induced by positioning reactants adjacent to each other
(2) diffusion accounted for the remaining interfacial mixing
(3) the collagen materials, so formed vary from those prepared in ground control experiments.
No further information discussing the experimental results could be located at this time.
This investigation had commercial applications in the area of biofilms which are used for artificial skin implants, blood vessels, tendons, corneas, and other organs.
(2) Wessling, F. C. and Maybee, G. W.: Consort 1 Sounding Rocket Flight. Journal of Spacecraft and Rockets, Vol. 26, No. 5, September-October 1989, pp. 343-351. (preflight; brief description of MDA)
(3) Wessling, F. C., Lundquist, C. A., and Maybee, G. W.: Consort 1 Flight Results - A Synopsis. Presented at the IAF 40th International Astronautical Congress, October 7-13, 1989, Mˆlaga, Spain, IAF #89-439. (post-flight)
(4) Cassanto, J. M.: Overview/Summary of Results from the Material Dispersion Apparatus (MDA) on Consort 1 Rocket Flight. Presentation to the Spring CMDS Scientific and Technical Project Review, Guntersville, Alabama, April 18-20, 1989. (post-flight)
(5) Input received from Principal Investigator M. Luttges, December 1991.