CA1080123A - Particulate material for use in the analysis of liquid samples - Google Patents

Abstract

ABSTRACT OF THE DISCLOSURE

A particulate material suitable for automatic assay of fluid samples, particularly immunossays, in which a reaction product of the assay is bound to the material and separated from the reaction mixture by use of a magnetic trap. The particulate material comprises a reagent and one or more magnetic particles in a matrix of binding material and has a controlled specific gravity approaching that of the fluid sample so as to retard separation of the particulate material when mixed with the fluid sample. The binding material is selected from a group consisting of cellulose or a cellulose derivative, a polymer or synthetic polymeric material, and agarose. The material is of especial use in radioimmunoassays carried out in an automated continuous-flow manner.

Description

1(~801Z;~
This invention is concerned with a particulate material useful for analysing a fluid sample, particularly but not exclusively for use in immunoassays.
This application is a divisional application of applicant's copending application Serial No, 273,631, filed March 10, 1977.
It is known to assay biological fluids such as blood serum or urine to detect and quantify the presence therein of antibodies (and similar binding proteins), antigens (and similar substances such as haptens) and antibody-antigen immune complexes. Such procedures are broadly called immunoassays. One common technique in immunoassays is to make use of the binding reaction which takes place between a limited amount of an antibody and two antigens, both antigens being capable of binding ;~
with the antibody but being distinguishable, e.g. in that one antigen carries an identifying label. The proportion of labelled antigen which binds with the antibody gives an indication of the amount of unlabelled antigen present. Thus, if for example a biological fluid sample containing a specific antigen is mixed with antibody and an amount of labelled antigen, the amount of (unlabelled) antigen in the sample can be determined.
Various labels have been suggested, but the most successful has been a radioactive label and immunoassays using radioactive labels are called "radioimmunoassays" (RIA).
In immunoassay procedures, it is often necessary to separate the reaction product (e.g. the antibody-antigen complex) from the raction mixture in order for example to determine the amount of labelled antigen in mb/l D - 1 ~

1080~Z3 the product (by direct analysis either of the product or of the remaining reaction mixture). It has been suggested in the prior art that in RIA procedures (in which a separation step is essential), the antibody be immobilised on a carrier to facilitate subsequent separation of the reaction product. Thus, in "Continuous Flow Automated Radioimmunoassay Using Antibodies Attached to Red Blood Cells", by S. J, Luner, Analytical Biochemistry, Vol, 65 (1975), pages 355-364, it is said that red blood cells may be used for the immobilisation of antibodies in automated RIA techniques using a continuous-flow system.
In "Magnetic Solid-Phase Radioimmunoassay" by L. S. Hersh and S. Yaverbaum, Clinica Chemica Acta, Vol. 63 (1975), pages 69-72, antibody is immobilised on magnetic particles and the reaction product separated from the reaction mixture by application of a magnetic field. This RIA proeedure is carried out in a test tube reaetor, and the supernatant reaetion mixture is assayed.
We have now devised a partieulate material `
useful for this purpose, by whieh a series of liquid samples ean be sueeessively and automatically assayed by, for exampleS RIA.
The invention relates to a particulate material for use in the analysis of a liquid sample of known speeifie gravity. The particulate material comprises a resgent and one or more magnetic particles in a matrix of binding material The particulate material has a controlled specific gravity approaching that of the liquid sample so as to retard separation of the particulate material when mixed with the liquid sample.

mb/J a The binding material is selected from a group consisting of: cellulose or a cellulose derivative, a polymer or synthetic polymeric material, and agarose.
In use a reaction mixture is flowed along a conduit. The mixture comprises the sample under test which includes the constituent of interest, i e, the substance which is to be assayed. In the case, for example, of biological fluids such as blood serum, the constituent of interest may, for example, be an antigen, e.g. a peptide hormone, a steroid hormone, a drug or a virus (the term "antigen" is intended to include haptens and other similar substances), an antibody (which term includes other binding substances), or an antibody; antigen complex, or a single protein.
The invention is not limited, however, to the assay of biological fluids.
The reaction mixture also includes, as a solid phase, magnetically attractable particulate material which has a reagent bound thereto, This particulate material may itself be a composite material made up of, for example, a matrix containing magnetically attractable material, with the reagent bound to the particle.

mb/J ~ _ 3 _ , The magnetically attractable material may be, for example, iron or magnetic iron oxides, nickel, cobalt or chromium oxide.
, Suitably, one or more particles of such a material are ¦ 5 embedded in the matrix. The matrix itself may be of a wide variety of materials including many synthetic and natural polymeric materials (e.g. cellulose, cellulose , derivatives, agarose, organic polymers). The reagent ;~
may be bound directly to the matrix or to another material within the matrix.
The reagent itself is a substance which takes part in a reaction in the reaction mixt1lrc. It may react directly with the constituent of interest in the sample, or it may react not directly with the sample but with a second reagent in the mixture. Thus, for example, the reagent on the particulate matter may be an antibody which will react directly with an antigen in the sample under test, or the sample itself may contain an antibody and the reagent react with an antigen added as the second reagent. In the latter case, the second reagent antigen may also react with the antibody in the sample.

1080iZ3 :

The reagent is bound to the particulate -~
material in such a manner that the reagent is avallable to react with another substance in the reaction mixture.
Usually, there will be reagent bound to the periphcral surface of the particulate material, but this is not essential provided that the reagent is accessible for reaction. The reagent may thus be wholly within the matrix but in such case, the matrix will be porous to the liquid of the reaction mixture.
The nature of the reagent can vary very widely, depending on the particular analysis to be performed. It may, for example, be an immunoglobulin, an antigen (e.g.
a virus) or another biological substance. After reaction, it remains bound (as reaction product) to the particulate material and is thus separated with the particulate material from the reaction mixture.
The separation of the particulate material solid phase from the liquid phase of the reaction mixture is effected in the conduit using a magnetic trap. The reaction mixture flows along the conduit into the resion of a localised magnetic field (the magnetic trap), ~-here-upon the solid phase is held by the field whilst the liquid phase flows on.
Preferably, the magnetic field is disposed in the conduit substantially transversely of the flo~-ing reaction mixture, but this is not essential. As ~ill be clear, the strength of the field must be sufficient to hold the solid phase against the liquid flow.
After the liquid has passed throush the trap, the solid phase in the trap in the conduit can be washed by passing a wash liquid through that portion of the conduit. The solid phase remains held in the trap but the particles are exposed to, and washed by, the wash liquid flowing past. It is a Highly advantageous pre-ferred feature to be able to wash the solid phase "on line" and thus the apparatus preferably comprises means for passing wash liquid along the conduit.
The wash liquid may be water or any inert fluid or solution, its purpose being to remove from the solid phase remaining traces of the reaction mixture liquid phase. This is particularly important in immuno-assays, such as RIA, where the solid phase is to be assayed for the presence of a label, since in such cases even trace residues of the reaction mixture liquid phase could lead to incorrect assay results.
The magnetic field is preferably provided by at least one magnet means actuable to provide magnetic field in a portion of the conduit. The (or each) magnet means may be a permanent magnet which is mo~eable to vary the field from a minimum (when the trap is "de-actuated") to a maximum "when the trap is "actuated"). We prefer, however, to use one or more electromagnets. Upon de- -energisation of the electromagnet, it is preferable to degauss by providing an alternating current, thus ridding the electromagnet of residual magnetic field. Solid phase in the trap will also be demagnetised by this procedure, thus reducing any tendency to clogging due to magnetic attraction.
The magnetic trap may consist of a single 10801Z3 ~ ~

~agnet means, or two or more such means. For many pur-poses, it is preferred to provide two (or more~ magnetic traps, spaced apart along the conduit. This enables an improved washing procedure, for example, to be effected. ~-Thus, with two traps, the magnet means of each trap are actuable independently of each other, and the solid phase is separated from the liquid phase of the reaction mixture in the first (upstream) trap~ It is washed whilst it is held in that trap, and then the trap is de-actuated to release the solid phase into suspension in flowing wash liquid. The particulate matter is carried to the second trap (which is energised) and held against the wash liquid flow. This double wash procedure is ~
particularly effective.
The liquid phase of the reaction mixture passes through the magnetic trap(s) and flows further along the conduit. Similarly, after washing, the solid phase is released from the trap and passes along thc washed con-duit. Preferably, valving means are provided in the conduit for directing the separated solid phase or liquid phase to the analysis means as required. If the solid phase is to be analysed, the liquid phase of the reaction mixture can be collected in a receiver or passed to waste~
If the liquid phase is to be measured (possibly together with wash liquid), the solid phase may be passed to a receiver, and possibly (after treatment) re-uscd. It will usually be preferable to assay the solid phase because it can be automatically washed (as described above), whereas the liquid phase is likely to be bulky (with the wash liquid) and not so easily handled or anal~sed.

~080123 The constituent of interest in the sample is determined by analysis of the separated solid cr liquid reaction mixture phases. It will be appreciated that this determination may involve several analytical and/or calculation steps. Thus, in the case of RIA, analysis of the solid phase by counting the radioactivity, will reveal the amount of radioaFtive label in the solid phase. From this, and standard curves, it will be possible to deter-mine the amount of constituent of interest in the sample.
It is possible to effect further reactions on the separated solid or liquid phase, and to introduce further magnetically attractable particles for a subsequent separation step. This may be desirable, for example, when the reaction mixture contains an enzyme or co-enzyme. In such cases further magnetic traps may be provided in the conduit downstream of the trap(s) for effecting the first separation.

The whole or any part of the reaction mixture may be preformed before it is flowed along the conduit. It is usually preferred, however, to add one or more reagents to the flowing sample (or partly formed mixture) in the conduit. Thus, the apparatus preferably includes means for introducing the said particulate material and/or the second reagent into admixture with the flowing sample (or reaction mixture) in the conduit.
Intermixing of the sample and the reagents will take place in the conduit as the mixture flows (as is described more fully hereinafter).
Whilst we have specifically referred above to : . :

~.
the reagent bound to the particulate material, and to - the second reagent, it will be understood that there may also be other reagents present in the reaction mixture.
These other reagents may be added in admixture with the -particulate material or second reagent, or they may be pre-mixed with the sample, or they may be added individ-ually to the flowing sample (or reaction mixture) in the conduit. The nature of these reagents (if any are used will be determined by the nature of the assay being per-formed. The reaction mixture may also contain other sub-stances such as buffers.
In immunoassays involving binding between, for example, an antibody and an antigen, the second reagent will usually carry an identifying label such as a radio-active atom, a fluorescent group, an enzyme or co-enzyme or a chemiluminescent material. Thus, for example, when the biologica] fluid sample is to be assayed for an anti-gen, the reagent on the particulate material will be an antibody for binding with the antigen, and the second re-agent will be an antigen capable of binding with the anti-body and which also carries a label. In the alternative, the fluid sample may be assayed for antibody by providing an antigen on the particulate material and using a labelled antibody as the second reagent. As will be understood by those skilled in the art, analysis of the separated solid phase or liquid phase for the amount of label present will allow determina'ion of the amount of constituent of interest in the fluid sample.
The particular analysis to be carried ollt on the separated solid or liquid phases will depend on the , ' assay being effected and (where a labelled reagent is used) on the nature of the label. Thus, the apparatus may lnclude, for example, means for measuring the radioactivity, the colour or fluorescence of the separated phase or i~s enzymic activity. -~
It will be understood that it is not essential in the method of the invention to use a second or any reagent other than that bound to the particulate matter.
Thus, the method (and apparatus) can be used, for example, to separate a particular constituent from a fluid sample (by selectively binding it to the reagent on the par- ;
ticulate material) and then subsequently assaying the separated solid phase, In most immunoassay procedures, however, a second reagent is used (and there may often be other reagents too).
The method and apparatus may be used in the known continuous-flow type of procedure in which individual segments of reaction mixture are passed along the conduit, separated by an inert fluid ~egment (e.g. air) and, if desired, a wash liquid seg-ment. This is described in U.S. specification no.

2,797,149 to which reference should be made for further details. Thus, the apparatus preferably includes means for passing successive reactio~ mixtures along said conduit separated from each other by at least an inert fluid segment of sufficient volume to occlude said cond~lit and maintain said successive mixtures dis-crete. The apparatus may also include means for intro-ducing inert fluid segments into said conduit to sub-divide li~uid samples or reaction mixtures thercin.

- . ...

Further, the apparatus preferably includes means for providing a wash liquid segment between successive reaction mixtures flowing in said conduit, each said mixture being separated from adjacent wash liquid seg-ments by at least an inert fluid segment.
When the reaction mixture (or sample) is segmented, and the particulate material and/or the second reagent is to be added to the flowing segments in the conduit, then means are provided for introducing the particulate material (and/or the second reagen~) on an intermittent basis so that they merge with successi~re sample segments in the conduit. Preferably, the inter-mittent introducing means includes means for returning particulate material not admitted to the conduit, to a reservoir therefor. Preferably, also, the apparatus includes means for introducing buffer solution inio said conduit alternately with said particulato material, so as (in use) to maintain the flow along said conduit sub-stantially constant.
It will be appreciated that it is important that any labelled reagent (or other reagent of critical importance in the final determination procedllrc) be added only to the sample or reaction mixture, and not to any t wash liquid or other segments in the conduit, since other-wise an incorrect assay may result.
It is convenient, particularly when segmented flow is used, to provide in the apparatus upstream of the magnetic trap(s) a sensing means to detect the passage of a reaction mixture along the conduit and to provide means for actuating the magnetic trap to trap the `` 108~1Z3 particles in that reaction mixture, in response to the ~ -sensing means. Depending on how far upstream of the trap the sensing means is located, a time delay may be needed before actuat~on.
The particulate matter will preferably have a specific gravity close to that of the liquid phase of ~-the reaction mixture, so that it does not tend to settle out (nor to float to the top) but rather remains in good admixture in suspension, Generally, the particulate matter will have a specific gravity substantially in the range Gf about 1.4 to 3.2, although values outside this range can be used.
It will be understood that, in the method ~and in use of the apparatus) sufficient time must be allowed after forming the reaction mixture for the desired reaction(s) to take place, before the solid phase is separated from the liquid phase To provide this period (commonly called "incubation"), the conduit may include, for example, an incubation coil, In order that the invention may be more fully understood, one embodiment setting out method and apparatus for using the particulate material will now be described, by way of example only, with reference to the accompanying drawings, in which:
FIGURE 1 is a flow diagram of one form of apparatus according to the present invention;
FIGURE 2 illustrates the composition of the fluid stream passed along the system of Figure l;
FIGURE 2A illustrates the two-phase laminar flow pattern within the fluid samples of Figure 2; and FIGURES 3A-3C illustrate the operation of the on-line magnetic traps to effect separation and working of the solid phase.
Referring now to the drawings, there is shown mb~ - 12 --., - . . ~ :

108VlZ3 a sampler arrangement 10 for supplying a series of liquid samples along a compressible pump tube 12. Sampler arrangement 10 may be of the type shown and described in U.S. Patent 3, o38,340. Pump tube 12 is incorporated in a peristaltic-type pump 14, of the type described in U.S. Patent 2,935,028. (which also controls tubes 38, 40,

3~, 34 l, 30, 30 ~, 32, 36, 52, 57, 84 and 80 as herein-after described). Probe 15 is controlled to be immersed alternately in successive sample receptacles 16, and a wash receptacle 18. As the probe 14 aspirates air between immersion into successive sample receptacles 16 and wash reservoir lo, the sample stream directed along pump tube 12 comprises successive samples, each separated from adjacent samples by air-wash liquid-air segments.
Accordingly, simple integrity is maintained during flow along the entire system, as hereinafter described. In addition to sampler 10, a source 20 of antibodies immobilized on the surface of magnetically attractable particles in suspension, an associated source 22 of the ' buffer solution, a source 24 of a labelled antigen in solution, and an associated source 26 of buffer, are pro-vided. The system is operated such that the solid phase from source 20, appropriately buffered, and thc lebelled antigen from source 24, appropriately buffered, are intro-duced in controlled discrete volumes and proper phases, to be mixed only with successive samples pumped along pump tube 12 and conduit 28. To this end, pump 14 com-prises a plurality of pump tubes, 30, 32, 34 and 36 having respective inlets in fluid communication with source 20 of the solid phase, source 2~ of the labelled 10801Z3 : ~

antigen and the associated buffer sources 22 and 26.
Also~ pump tubes 38 and 40 are provided for periodically injecting air bubbles to segment the fluid streams passed along pump tubes 30 and 34, to provide discrete segments of uniform concentration of the solid phase and maintain a constant flow rate through the system when valves 40 and 42 are operated. Each of the pump tubes 30, 32, 34 and 36 has an associated return conduit 30 ~, 32 l ,34 ~ and 36 ~, respectively, whereby fluid is re-circulated back to the respective sources 20, 2~, 22 and 26. The outlets of pump tubes 30, 32, 34 and 36 are connected to the inlets of three port-two-position valves, 42. 44, 40 and 46, respectively. One outlet of each such valve is connected to the assoclated return conduit. The remaining outlets of valves 40 and 42, associated with the solid phase, are multipled and connected along a mixing coil 48 to conduit 28 a-t junction A. Also, the remaining outlets of valves 44 and 46 are multipled and connected along conduit 50 to conduit 28 at junction B. Additionally, the outlet of pump tube 52, having an inlet exposed to air, is connected to segment the liquid stream along conduit 50 prior to introduction into conduit 28. As peristaltic pump 14 opcrates con-tinuously, air segments are continuously injected via air pump tubes 38, 40 and 52 so as to achieve both intra-sample segmentation and intra-wash liquid scgmentation as hereinafter described, whereby effective internal mixing of the individual segments of the flowing stream along conduit 28 is obtained.
Valves 40, 42, 44 and 46 are controlled by a ,: - - : - . , .

~0801Z3 pro~rammer 152, which is timed with respect to sampler 10, such that the buffered labelled-antigen stream along conduit 50, and.the buffered solid phase along mixing coil 48, are intrQduced in discrete volumes and in pro-per phase at junctions B and A, respectively, to be inter-mixed only with each liquid sample along conduit 28.
During passage of the wash liquid segment (intermediate successive samples) along junction A and also along junction B, valves 42 and 44 are operated to position I, to interconnect pump tubes 30 and 32 with conduits 30' and 32', respectively, and recirculate the liquids being passed therealong to their sources. Simultaneously, valves 40 and 46 are operated to position II, to direct the buffer solution from sources 22 and 26, respecti~ely, to junctions A and B, to maintain constant flow rates along conduit 28. At this time, air i9 being pumped constantly along pump tube 52, wher~by air bubbles are periodically injected into conduit 50 to segment the fluid stream therealong and direct it to junction B,-to ensure proper mixing along coil 55. During passage of a sample along junctions A and B, programmer 152 operates valves 42 and 44 momentarily to position II, to introduce controlled discrete volumes of the buffered solid phase along mixing coil 48 and of the labelled antig~n sus-pension along conduit 50 into such sample. A m;~ing coil 55 is provided along conduit 28 and intcrmediate junctions A and B, to ensure that tne labelled antigens are thoroughly mixed and uniformly distributed through-out each liquid sample segment, prior to introduction of the solid phase at junction A.

The successive sample segments, co-mixed with the solid phase and labelled antigen to form reaction mixture, are directed along conduit 28 to an incubation coil 54. The composition of the stream directed to the incubation 54 is illustrated in Figure 2. Each of the segments~ or aliquots, of samples 51' S2, etc. include controlled volumes of the solid phase and labelled antigen; the wash liquid segments (W) are between successive samples and the successive samples are themselves segmented by air bubbles introduced along pump tube 52. In Figure 2A, the labelled antigen and the solid phase, i.e. antibodies immobilized on ma~net-ically attractable particles, are distributed throughout a sample aliquot and are indicated by "-" and "~", res-pectively. The presence of air segment immediately pre-ceding and following a liquid segment induces a two-phase laminar flow pattern, whereby tho liq~lid in such segment i9 caused to recirculate, as indicated by the arrows in Figure 2A. Such flow pattern results, primarily, from the drag imposed on the moving liquid immediately at the inner surface of conduit 28, and serves to accelerate the mixing of the various componcnts of the reaction mixture.
As described, the solid phase comprises mag-netically attractable particles of controllcd spccific gravity. Preferably, these particles comprise ~ matrix having one or more small ferromagnetic particles embedded therein. For example, the ferromagnetic particles may be coated with an organic material, e~g~ a polymeric material, or they may bs silanized, to bind with either ~080123 an antibody or antigen. For example, sui*able coating techniques have been described in the abo~e-identified Hersh and Yaverbaum article and, also, in t'The Pro-perties of Magnetic Supports in Relation to Immobilized Enzyme Reactions", P.J. Robinson et al, Biotechnology and Bioengineering,,Vol.XV. (1973) pp. 603-606. The ratio of the respective volumes of the matrix and the ~-magnetic particles is chosen so that the resulting specific gravity approaches that of the liquid phase.
As such, these magnetically attractable particles are carried along by the two-phase laminar flo~r pattern induced within each sample aliquot and do not tend to settle out, either by gravitation or flotation. It should be appreciated that the specific gravi~y of the magnetic particles need not be equal to the specific gravity of the liquid phase, but should be such as to avoid settling or flotation of the solid phase during passage through the qystem. The reduced density of such particles pro~ides for excellent wash characteristics, whereby there is substantially no contamination between successive samples. For example, if the particles were to have a ~pecific gra~ity much higher than that of the liquid phase, they would tend to settle out and collect in the cusp defined at the interface between the sample aliquot and the up-stream air segment along the inner wall surface of conduit 28. Such settling would sub-stantially increase the possibility that such particles could pass under the up-stream air segment, and follow- -ing liquid and air segments in the flowing stream to contaminate a following sample. If the spccific gravity - , .. ..

of the particles is reduced much below that of the sample, the particles separate out by flotation and may possibly pass over the following liquid and air segments to effect contamination. By controlling the specific gravity of the solid phase, so that it is retained and carried within the two-phase laminar flow pattern, con-tamination between successive samples is avoided.
The reaction mixture stream, with the solid phase constantly circulating within the individual sample aliquots, is passed through an incubation coil 54 to enable the reaction to proceed. The output of incubation coil 54 is greatly diluted by buffer directed along conduit 53 and pump tube 57, to inhibit the reaction (i.e. quench the reaction) and facilitate washing of the solid phase by a dilution process. The output of incubation coil 54, is directed alons conduit 56 to a wash statiorL~ identified as 58. SUCh wash station pre-ferably inclucles a firgt magnetic trap 60 and a second magnetic trap 62, arranged with respect to conduit 56 in on-line fashion. Each of the magnetic traps 60 and 62 are electromagnetic and arranged to direct magnetic flux transversely to the path of the sample stream. The magnetic traps 60 and 62 are operated in particular timed sequence by programmer 64 controlled by a sample detector 65. Accordingly, sample detector 65 comprises light source 66 and a detector 68 and is located at the output of incubation coil 54. Light from source 66 is directed transversely through conduit 56. l~hen a sample passing along conduit 56 intercepts such light, at time t of Figure 2, the reduced output level of a detector 68 instructs programmer 64 to energize the magnetic traps 60 and 62 concurrently.
l~hen energized, first magnetic trap 60, in effect, sweeps the solid phase from each of the sample aliquots comprising, for example, sample S1 as sho~n in Figure 2. The intensity of the magnetic fields generated by the first magnetic trap 60 and, also, the second magnetic trap 62, as hereinafter described, are sufficient to ensure that passage of intra-sample air bubbles -.~ithin sample S do not dislodge or carry away any of the solid phase retained within the wash station 58. Accordingly, the solid phase in each sample, albeit carried by individual aliquots, is accumulated along conduit 56 passing through the first magnetic trap 60. Energization f magnetic trap 62 at time t1 ensures that any solid phase not swept by magnetic trap 60 is retained. ~agnetic trap 60 is energized during passage of the entire Yolume of sample S and, also, during passage of at least a portion of the following wash liquid segment, i.e. during time interval t1 - t~ shown in Figure 2. Passage of such wash liquid segment through wash station 58, while the solid phase is packed, serves to remove any super-natant from the magnetic particles. Before the entire wash liquid segment has been passed, at time t, pro-grammer 64 de-energizes magnetic trap 60. Accordingly, the solid phase is caused to be resuspended ~ithin the remaining wash liquid segment. Air bubbles in the wash liquid segment, as introduced along pump tubes 52, tend to break-up the solid phase packed by the magnetic trap 60 and accelerate resuspension of the same in the wash . . .
liquid to ensure complete removal of any liquid phase, ;~
i.e. unbound labelled or unlabelled antigens, from the surfaces thereof. As the second magnetic trap 62 is energized, the solid phase is again swept from the ~-ash liquid segment.
In a particularly preferred embodiment of the in~ention, conduit 56 passing through the wash arrange-ment 58 is connected to the inlet of a three-port two-position valve 70, the outputs of such valve being connected to a solid-phase scintillation counter 72 (position 1) and waste (position II), respectively.
Also, an additional three-port, two position valve 76 is provided, having an inlet connected along conduit 78 to pump tube 80, whose inlet is connected to a source 82 of buffer, and having outlets connected to waste W
(position I) and solid-phase counter 72 (position II).
Valves 70 and 76 are controlled simultaneously by pro-grammer 64. ~n additional air pump -tube 84, llaving its inlet exposed to air, is connected to conduit 78, so as to periodically introduce air bubbles into the buffer stream flowed along conduit 78. The buffer liquid pumped along tube 78 is used for cleaning the solid-phase counter 72. As known, the presence of air bubbles alons conduit 78 serves to accelerate the cleaning of any residues of a preceding sample within the solid-phase counter. When the solid phase in magnetic trap arrangement 58 has been thoroushly washed and prior to the appearance of the next successive sample adjacent the light source 66, at time t3 of Figure 2, magnetic trap 62 is de-energized and valves 70 and 72 are operated momentarily to position I, to direct th~ solid phass, now suspended in wash liquid, to solid-phase counter 72 and the segmented buffer stream along conduit 78 to waste. At this time, the solid phase is passed through the storage coil 79 of solid-phase counter 72 and its radioactivity measured. Such measure-ments can be recorded, for example, by a print-out recorder 80~ Subsequently, valves 70 and 72 are operated to position II, to direct the fluid stream along conduit 58 to waste and the segmented buffer stream alon~ conduit 78 through the solid-phase counter, preparatory to the washing and measurement of the sol.id-phase in a next sample.
It will be evid~ent that the liquid-phase can be measured, if desired, rather than the solid phase~ To this end, a liquid-phase scintillation counter could be substituted for the solid-phase counter 74. In such event, the magnetic traps 60 and 62 arc operated, as described, to sweep out the solid-phase from the aliquots of the same'~ample. Concurrentl~, valves 70 and 72 are controlled (to position I) to pass the separated liquid phase directly to the storage coil of the liquid-phase counter and while magnetic traps 60 and 62 arc energized.
Subsequently, valves 70 and 72 are operated to position II, prior to de-energization of magnetic traps 60 and G2, such that the solid-phase re-suspended in thc wash liquid is passed to waste and the segmented buffcr liquid is passed to wash the liquid-phase counter. In either event, it can be appreciated that the controlled specific gravity of the magnetic particles, on which the solid phase is immobilized, and the magnetic in-line trapping technique described, co-operate to achieve a positive separation of 1~801Z3 , the solid and liquid phases, without deteriorating the wash characteristics of the system.

The individual samples passed along the conduit 28 may be reacted selectively.
For example, we can provide a plurality of solid phase sources 20 and label sources 24, each having an assoc-iated yump tube 30 and return conduit 30', each of wllich can be connected to a respective three part, two-position changeover valve. The outputs of tlle change-over valves for each of the solid phase and label systems as well as the outputs of the buffer valves may be connected to corresponding inlets of a multi input/single output valve, whose output is connected to junctions A and B.
Each of the single output valves is corltrolled in response to information read frorn the individual receptacle 16, so as to introduce the appropriate solid phase and label phase, in phase with each other, at junctions B and A, so as to react selectively each sample.
Also, the output on recorder 88 may be appropriately identified.

Claims (6)

THE EMBODIMENTS OF THE INVENTION IN WHICH AN EXCLUSIVE
PROPERTY OR PRIVILEGE IS CLAIMED ARE DEFINED AS FOLLOWS:

1. A particulate material for use in the analysis of a liquid sample of known specific gravity, said particulate material comprising a reagent and one or more magnetic particles in a matrix of binding material, said particulate material having a controlled specific gravity approaching that of said liquid sample so as to retard separation of said particulate material when mixed with said liquid sample, said binding material being selected from a group consisting of:
cellulose or a cellulose derivative, a polymer or synthetic polymeric material, and agarose.

2. The particulate material of claim 1, wherein said specific gravity of said particulate material is approximately equal to 1,4.

3. The particulate material of claim 1, wherein the magnetic particles are selected from a group consisting of iron, magnetic iron oxides, nickel, cobalt or chromium oxide.

4. The particulate material of claim 1, wherein said reagent is selected from a group consisting of an antigen, an antibody or biological substance.

5. The particulate material of claim 1, wherein said reagent is an antigen and said liquid sample comprises an antibody for reaction with said antigen.

6. The particulate material of claim 1, wherein said reagent is an antibody and said liquid sample comprises an antigen for reaction with said antibody.