Capillary hematocrit separation structure and method

Details Capillary hematocrit separation structure and method Capillary hematocrit separation structure and method

1999-10-28

2001-11-20

Wallenhorst, Maureen M. (Department: 1743)

Chemistry: analytical and immunological testing

Sedimentation rate or hematocrit

C436S165000, C436S175000, C436S177000, C422S051000, C422S073000, C422S105000, C422S105000

Reexamination Certificate

active

06319719

ABSTRACT:

BACKGROUND OF THE INVENTION
The present invention is directed to physical structures and methods for separating hematocrit out of small volume whole blood samples leaving merely the plasma or plasma containing a substantially reduced partial volume of hematocrit. The present invention is particularly directed to such structures having no moving parts which subsequent to separation of the plasma would facilitate contacting the reduced hematocrit content plasma with a dry reagent to permit an accurate detection of an analyte.
Many diagnostic tests are carried out in the clinical field utilizing a blood sample. It is desirable, when possible, to use a very small volumes of blood, often no more than a drop or two. Capillary structures are often employed when handling such small volumes of blood or other fluids. The presence of the hematocrit in the blood sample often interferes with accurate testing and so the removal of, or reduction in concentration of, the hematocrit in the sample, leaving a reduced hematocrit content plasma for testing, is often desirable or even necessary. The removal of the hematocrit is often done using a filter. An example of such a filter device employing capillary structures is described in Hillman, et al., U.S. Pat. Nos. 4,753,776 and 5,135,719. Other devices employing capillary structures to handle whole blood samples are disclosed in McDonald, et al., U.S. Pat. No. 5,039,617; Hillman, et al., U.S. Pat. No. 4,963,498; and Columbus, U.S. Pat. No. 4,271,119.
While such filter devices generally perform satisfactorily, many filter materials tend to absorb a significant portion of the plasma from the blood sample thus leaving only a small volume of the reduced plasma for analytical testing. As the total volume of the sample is diminished, the proportion of the plasma fraction that is absorbed by the filter tends to increase leaving even smaller volumes for testing. It is therefore desirable to construct alternative means for removing hematocrit from whole blood that would be usable on very small sample volumes.
SUMMARY OF THE INVENTION
A capillary hematocrit separation structure according to the present invention is included within a housing having a fluid inlet port, a vented reaction region, and a capillary pathway connecting the inlet port and the reaction region. The capillary pathway is dimensioned so that the driving force for the movement of liquid through the capillary pathway arises from capillary pressure. A plurality of obstructions are fixed in the capillary pathway, each obstruction having a concave portion facing toward the vented reaction region. The situation of the concave portion is to be understood, in reference to the flow of liquid from the fluid inlet port to the reaction region, to be positioned on the down stream side of the obstructions.
The obstructions can take on a variety of shapes including a bullet shape and a quarter moon shape, a ¾ quarter moon shape being preferred. The obstructions are to be situated far enough from each other so that their mere proximity to each other does not create a filter effect, yet they are to be situated close enough to each other as to minimize the volume of liquid retained in the capillary pathway. Preferably, the obstructions are separated from each other, on a nearest neighbor basis, by about 10
−5
meters, and arranged in a hexagonal close-pack configuration.
The number of obstructions to be employed is determined by the capacity of the concave portions of the obstructions. It has been observed that as whole blood flows through a capillary pathway containing a plurality of obstructions in accordance with the present invention, hematocrit collects in the concave portions of the obstructions. While it was initially thought that the hematocrit collected by virtue of Von Karmen vortices, it has now been determined that such vortices only occur in turbulent flow circumstances, and the passage of blood through a capillary channel is probably laminar. The mechanism behind this effect has not been identified, but the effect is significant enough to permit substantial reduction in the partial volume of hematocrit in whole blood samples.
It is desirable that sufficient obstructions are employed to provide concave portions having a total volume exceeding the volume of hematocrit in the expected sample size. In a preferred embodiment the volume of each concave portion is about 10
−13
to 10
−14
m
3
, or 10
−4
to 10
−5
&mgr;l. A sample comprising a single drop of blood typically has a volume of 20 to 50 &mgr;l., of which typically 35% to 45% constitutes hematocrit. Even smaller volumes of whole blood are often used by diabetics and others during testing, the smaller volume being achieved by expressing the blood sample from a small cut or puncture, in which case the volume of the sample may amount to only 2 to 10 &mgr;l. A capillary hematocrit separation structure according to the present invention capable of separating the hematocrit from a single drop sample size includes about 10
4
to 10
5
obstructions. A capillary pathway in such a structure can be a rectangular channel about 100 &mgr;m high or less, 2 to 5 mm wide, and up to 70 mm long. The volume of the inlet port leading to the capillary pathway is generally less than about 50 &mgr;l, and preferably the sum of the inlet port volume and the capillary pathway volume is less than about 20 &mgr;l.
It is to be understood that the number of capillary pathways between the fluid inlet port and the reaction region is not critical and that one or more than one can be employed, if desirable, to facilitate to the construction of the obstructions or other features of the device. While the hematocrit is observed to preferentially accumulate in the concave portions, it is also observed to accumulate to a lesser extent in other regions of the structure, particularly adjacent to the walls defining the capillary pathway.
A capillary hematocrit separation structure according to the present invention can be molded as two pieces of a thermoplastic resin such as nylon, styrene-acrylic copolymer, polystyrene, or polycarbonate using known micro-injection molding processes. The mold for making the obstructions in the capillary pathway can be constructed by deep reactive ion etching processes typically employed in the manufacture of molds for pre-recorded compact disks and digital video disks. A reaction region is generally also formed by the same process at an outlet end of the capillary pathway which is generally vented to ensure that there is no opposition to the fast capillary flow of liquid through the capillary pathway. The capillary pathway and the reaction region in the molded structure is then preferably subjected to a hydrophilizing process such as by plasma etching or DONS solution. A suitable dry reagent can be situated in the reaction region, if desired. The pieces of the structure are then assembled so that the capillary pathway and reaction region are enclosed within the structure, yet can be accessed at an inlet port designed to receive a sample of blood.
The resulting structure can be viewed as an apparatus for separating hematocrit from a whole blood sample having a selected total volume, the sample including a partial volume of blood plasma and a partial volume of hematocrit. The fundamental features of the apparatus comprise a body having an inlet port for receiving a whole blood sample, a vented reaction region spaced from the inlet port, and at least one capillary pathway having an inlet end coupled to the inlet port and an outlet end coupled to the vented reaction region, each capillary pathway being dimensioned sufficiently small to assure transport of blood plasma from the inlet end to the outlet end by capillary pressure delivering a reaction volume of plasma to the reaction region, each capillary pathway including a plurality of obstructions, each of the obstructions having a concave portion facing toward the outlet end of the pathway, the sum of the concave portions having sufficient volume to contain at l

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