Classifications

• • B—PERFORMING OPERATIONS; TRANSPORTING
  • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
  • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
  • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
  • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
  • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
  • B01L3/5021—Test tubes specially adapted for centrifugation purposes

Definitions

• the present invention is directed to ultracen- 5 trifuge tubes and, more particularly, is directed to a multi-chamber ultracentrifuge tube.
• VLDL very low density lipoproteins
• LDL low density lipoproteins
• HDL high density lipoproteins
• lipoprotein sec ⁇ gation involves the utilization of a preparative ultra ⁇ centrifuge after which the separated lipoproteins are subjected to analytical measurements to determine the cholesterol concentration. Since the sample is normally placed in a typical single chamber centrifuge tube for insertion in a fixed angle or vertical tube ultracentri ⁇ fuge rotor, the separate bands formed during centrifuga ⁇ tion will reorient after the centrifugation is com ⁇ pleted. Some residue from one band may be left on the tube wall and contact other bands as they reorient to a new position affecting the purity of the separation.
• One type of single chamber tube used is that which is dis ⁇ closed in U.S. Patent 4,301,963 issued to Steven T. Nielsen on November 24, 1981 and entitled "Integral One piece Centrifuge Tube.”
• each separated lipoprotein fraction must be physically isolated so that it is not contaminated and will not result in an incorrect reading of the cholesterol concentration for that particular density lipoprotein.
• Typical techniques utilized to accomplish this separation are centrifuge tube slicing or aspiration. In the tube slicing technique the cutting blade must be carefully positioned and be maintained in an extremely clean condition. Furthermore, depending upon the particular type of tube utilized, a significant amount of force may have to be applied by the user to pierce the tube. Once the tube is sliced, the upper portion of the tube is isolated and the separated lipo- protein can be aspirated out of the sliced-off portion of the tube without disturbing the other separated lipopro ⁇ tein bands in the remainder of the tube.
• bands are pipetted or suctioned off layer by layer. This operation may be simple and relatively convenient, but it poses a strong possibility of contamination. This is because lipoproteins from the removed fraction may adhere to the side of the tube and contaminate the remaining fraction.
• the present invention is directed to a multi- chamber ultracentrifuge tube.
• the separate chambers in the centrifuge tube are joined by a constricted area which forms, a conduit to provide fluid communication between the chambers during centrifugation of the tube.
• a fluid sample placed into the centrifuge tube will, during centrifugation, be separated into lighter material in the chamber closest to the rotational axis of the rotor, while the heavier material will be separated into the chamber farthest from the spin axis of the rotor.
• a support spacer is placed adjacent the constriction to provide support to the tube at the con ⁇ stricted area during centrifugation.
• constricted area Various configurations of the constricted area are applicable to the present invention. Further, the number of separate chambers that can be formed is limited only by the length of the tube.
• Use of the present invention allows an inves ⁇ tigator of blood serum lipoproteins to accomplish an efficient separation of the various density lipoproteins without contamination. Subsequent to the centrifugation run, the constricted area of the tube can be severed and sealed to ensure the isolation of the different density lipoproteins.
• Figure 1 is an elevational view of the ultra ⁇ centrifuge tube of the present invention
• Figure 2 is an elevation sectional view of the ultracentrifuge with an annular support spacer and a top support spacer;
• Figure 3 is a top view of the split annular spacer of Figure 2;
• Figure 4 is a sectional view of the annular spacer taken along the lines 4-4 in Figure 3;
• Figure 5 is a partial sectional view of a cen ⁇ trifuge rotor with the present invention in place, show ⁇ ing the orientation of the various separated fluid sample bands during centrifugation;
• Figure 6 shows the reorientation of the fluid sample bands in Figure 5 when the tube is at rest in a vertical position
• Figure 7 shows an alternate embodiment of the present invention utilizing a wedge spacer in a centri ⁇ fuge rotor with the wedge facing the spin axis of the rotor;
• Figure 8 is similar to the centrifuge tube shown in Figure 7 with the wedge-shaped spacer facing away from the spin axis of the rotor;
• Figure 9 is a second alternate embodiment of the present invention showing several chambers in the centrifuge tube with several annular support spacers.
• Figure 10 is a third alternate embodiment of the present invention showing several chambers within the centrifuge tube separated by multiple wedge spacers.
• FIG. 1 A multi-chamber ultracentrifuge tube 10 of the present invention is shown in Figure 1 having a generally uniform cylindrical wall 12 enclosed with a hemispherical bottom 14 and a bell-shaped upper portion 16. Located between the upper portion 16 and the hemispherical bottom end 14 is a constricted area 20 forming an interior con- duit 22 that provides fluid communication between an upper chamber 24 and a lower chamber 26. Positioned on the upper portion 16 is a fill port 28 which is open (shown in phantom) for receipt of a fluid sample prior to centrifugation. After the tube is filled with the sample to be centrifugated, the fill port 28 is hermetically sealed to form a solid seal 30.
• the tube 10 is prefera ⁇ bly an integral one-piece tube of a thin-wall polyallomer of the type explained more fully in the above-referenced U.S. Patent 4,301,963.
• the centrifuge tube 10 is designed so that its constricted area 20, having the shape generally of an hour glass, compatibly receives an annular support spacer 32.
• the diameter of the outside surface 34 of the annular spacer is approximately the same as the diameter of the cylindrical wall 12 of the tube.
• the spacer 32 provides support to the thin-wall polyallomer tube 10 during centrifugation, so that there is no distortion of the tube or constriction of the con- duit 22 which would affect fluid communication between the upper chamber 24 and the lower chamber 26.
• the configuration of the spacer 32 is shown in Figure 3 with two half sections 38 and 40 that form the entire circular arrangement of the split annular spa ⁇ cer.
• the central opening 41 formed by the spacer sec- tions 38 and 40 is approximately the same size as the exterior diameter of the constricted area 20 on the tube 10 in Figure 1.
• the sectional configuration of one por ⁇ tion 38 of the split annular spacer 32 is shown in Figure 4.
• the sectional cross area 42 is designed to occupy and conform to the constricted area 20 in the tube 10.
• centrifuge tube 10 in Figure 1 have two chambers 24 and 26 with a constriction 20 between the chambers that forms a conduit 22 which will be large enough to allow fluid communication between the cham- bers.
• the upper chamber volume should be approximately 31% of the total tube volume and the lower portion be 69% of the total tube volume.
• This chosen relative volume for each of the upper and lower chambers is for analysis of a serum sample to ensure that during centrifugation the very low density lipoprotein fraction will remain in the upper chamber after centrifugation, while the combination low density and high density lipo ⁇ protein fraction will remain in the lower chamber.
• the fill port 28 in Figure 1 is sealed.
• the tube 10 is then partially lowered into a tube cavity 44 in the rotor 46 in Figure 5.
• the split annular spacer 32 is placed around the constricted area 20 of the tube 10 and the tube is lowered completely into the cavity 44 as shown in Figure 5.
• the top floating spacer 36 is positioned above the tube in contact with its upper por ⁇ tion 16.
• the rotor lid 48 is placed on the rotor and the centrifugation process is initiated.
• the very low density lipoproteins 50 will band in an area of the tube closest to the spin axis 52 of the rotor 46.
• the low density and high density lipoprotein portion 54 will form a band in the portion of the tube farthest from the spin axis 52 of the rotor.
• the respective upper and lower chambers are sized so that the entire very low density lipopro ⁇ teins 50 will remain in the upper chamber 24, while the low density and high density lipoproteins will remain in the lower chamber 26.
• the lipoprotein fractions 50 and 54 will be separated by an inner band of the blood sample 56 that does not contain any lipoproteins.
• tube 10 is removed from the centrifuge rotor and the constricted area 20 of the tube is pinched with a clamp, hemastats or possibly heated hemastats to seal off the conduit 22 and maintain the isolation of the separated lipoprotein con ⁇ tents in the respective chambers.
• the constricted area 20 may also be pinched off by twisting one chamber 360 ⁇ along its longitudinal axis with respect to the other chamber.
• the contents from the chamber are transferred to another tube for the cholesterol analysis. There is no requirement of tube slicing or aspiration to separate the lipoprotein bands.
• the content of the lower chamber 26 which con- tains both the low density lipoproteins and high density lipoproteins is transferred to another two-chamber tube 10 and the centrifugation process is repeated so that the lighter low density lipoprotein fraction will flow to the top of the upper chamber 24 and the heavy density lipo- protein will be in the lower chamber 26.
• the constricted area 20 is pinched off to seal the conduit 22 and maintain the separation of the respec ⁇ tive lipoprotein constituents.
• ultracentrifuge tube 60 has an upper chamber 62 and a lower chamber 64 which are joined by a restricted area 66 forming a conduit 68. While the embodiment of the pres ⁇ ent invention shown in Figure 1 has the conduit formed symmetrically around the longitudinal center of the tube 10, the embodiment shown in Figure 7 has the conduit portion 68 closely adjacent the cylindrical wall 70 of the tube 60. The bottom hemispherical portion 72 of the tube 60 and its upper bell-shaped upper portion 74 estab- lish an enclosed centrifuge tube. A wedge-shaped spacer 76 is placed in the constricted area 66 to form exterior support to the upper chamber 62 and the lower chamber 64 during centrifugation.
• the orientation of the tube 60 in Figure 7 positions the conduit 68 so that it is as far away from the spin axis 52 of the rotor as possible. In this orientation the heavier components will move outward and collect on the straight interior wall 73 of the cen ⁇ trifuge tube and move smoothly into the lower chamber 64. However, the lighter material must follow a more oblique path around the wedge-shaped surface 78 of the constricted portion 66.
• FIG 8 the same centrifuge tube 60 is shown rotated 180° in the tube cavity 44 of the rotor 46 as compared to the tube position in Figure 7.
• the constricted portion 66 is oriented in a position as close as possible to the spin axis 52 of the rotor.
• This orientation of the centrifuge tube 60 has the advantage that the light fractions will collect on the straight surface 73 of the tube closest to the spin axis of the rotor and move smoothly up into the upper chamber 62.
• the constricted wedge-shaped por ⁇ tion 78 on the interior of the tube will result in a more oblique or circuitous path for the heavier fractions to move down into the lower chamber 64 which is farther away from the spin axis of the rotor.
• each are preferably made of a polyphenylene oxide material known as Noryl developed by General Electric. This material is advantageous, because its density is approximately 1.06 grams per ml which is closely approxi-
• FIG. 9 showing a second alternate embodiment of the present invention wherein the centrifuge tube 80 has four chambers 82, 84, 86, and
• a split ring spacer 34 as is shown used with the centrifuge tube 10 in Figure 2. In particular applications depending upon the size of the centrifuge tube and the size of the respective chambers 82, 84, 86, and 88, it is possible to facilitate separa- tion of the bands of a fluid sample into three or four zones.
• the centri ⁇ fuge tube 110 is shown having a plurality of chambers 112, 114, 116, and 118 which are separated respectively by constricted areas 120, 122, and 124.
• Each of the respective chambers 112, 114, 116, and 118 are in fluid communication with each other through the respective conduits 126, 128, and 130.
• the respective restricted areas 120, 122, and 124 receive wedge spacers 76 similar to that arrangement as shown in conjunction with the
• the tube of the present invention is preferably made of a polyallomer material because of its density proximity to blood serum, it is possible to consider other thermoplastic polymers such as thermoplastic polyester, polypropylene and poly ⁇ ethylene.
• centrifuge tubes other than those specifically shown in this appli ⁇ cation could be designed within the scope and spirit of the present invention.

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• 1983
  • 1983-05-16 WO PCT/US1983/000771 patent/WO1984000313A1/en not_active Application Discontinuation
  • 1983-05-16 EP EP19830902064 patent/EP0112868A1/de not_active Withdrawn
  • 1983-05-16 JP JP1984600003U patent/JPS59500008U/ja active Pending

Non-Patent Citations (1)

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