U.S. patent application number 14/611443 was filed with the patent office on 2015-06-18 for sample collection and analysis.
The applicant listed for this patent is IDEXX Laboratories, Inc.. Invention is credited to Matthew Howard Myles.
Application Number | 20150168404 14/611443 |
Document ID | / |
Family ID | 51488262 |
Filed Date | 2015-06-18 |
United States Patent
Application |
20150168404 |
Kind Code |
A1 |
Myles; Matthew Howard |
June 18, 2015 |
Sample Collection and Analysis
Abstract
Management of the health status of an animal colony using a
plurality of blood collection cards and the analysis of dried blood
from members of the colony that has been collected on the cards.
Members of the colony may be removed from the colony as a result of
the analysis.
Inventors: |
Myles; Matthew Howard;
(Moberly, MO) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
IDEXX Laboratories, Inc. |
Westbrook |
ME |
US |
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|
Family ID: |
51488262 |
Appl. No.: |
14/611443 |
Filed: |
February 2, 2015 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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14538381 |
Nov 11, 2014 |
8945945 |
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14611443 |
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14089103 |
Nov 25, 2013 |
8927298 |
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14538381 |
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61776560 |
Mar 11, 2013 |
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Current U.S.
Class: |
506/9 ; 435/5;
435/7.1 |
Current CPC
Class: |
G01N 2469/10 20130101;
G01N 33/56983 20130101; G01N 1/30 20130101; G01N 33/5695 20130101;
G01N 33/569 20130101; G01N 33/56911 20130101; Y10S 436/811
20130101 |
International
Class: |
G01N 33/569 20060101
G01N033/569 |
Claims
1. A method of determining a presence or absence of an infectious
disease in a population of rodents, the method comprising: (a)
receiving a plurality of blood collection cards from a user
responsible for a population of rodents, wherein the blood
collection cards have at least one spot of dried rodent blood; (b)
extracting the blood from the cards; (c) conducting an immunoassay
for analyzing the extracted blood for a presence or absence of at
least one antibody for an infectious agent indicative of an
infectious disease, thereby determining the presence or absence of
the infectious disease in the rodent population; and (d) reporting
the results of the presence or absence of the infectious disease to
the user.
2. The method of claim 1, wherein the immunoassay comprises
contacting the extracted blood with a fluorescently labeled binding
partner for the antibody.
3. The method of claim 1, wherein the immunoassay is a multiplex
fluorescence immunoassay.
4. The method of claim 1, where in the extracting comprises
extracting the blood with a buffer comprising
ethylenediaminetetraacetic acid (EDTA).
5. The method of claim 4, wherein the immunoassay is a multiplex
fluorescence immunoassay.
6. The method of claim 1, wherein the members of the population of
rodents are mice.
7. The method of claim 1, wherein the members of the population of
rodents are rats.
8. The method of claim 1, wherein the blood collection cards each
have at least one collection area having an absorbent material
suitable for holding about 10-40 .mu.L of whole blood.
9. The method of claim 1, wherein the blood collection cards each
have at least two collection areas.
10. The method of claim 9, wherein the at least two collection
areas are separated by a perforation that allows separation of the
cards such that a single sample area is on each separated card.
11. The method of claim 6, wherein the step of analyzing the
extracted blood comprises analyzing for the presence or absence of
seven or more diseases selected from the group consisting of: a.
MHV; b. MVN (MMV); c. NS1 (Generic Parvovirus); d. MPV (MPV1-5); e.
MNV; f. TMEV; g. EDIM; h. Sendai virus; i. Mycoplasma pulmonis; j.
PVM; k. REO3; l. LCMV; m. Ectromelia virus; n. MAD1; o. MAD2; p.
Polyoma virus; q. Encephalitozoon cuniculi; r. CARB; s. Clostridium
piliforme; t. MCMV; u. K virus; v. Hantaan virus; w. Lactate
dehydrogenase-elevating virus; and x. MTV.
12. The method of claim 7, wherein the step of analyzing the
extracted blood comprises analyzing for the presence or absence of
seven or more diseases selected from the group consisting of: a.
RCV; b. NS1 (Generic Parvovirus); c. RPV; d. RMV; e. KRV; f. H-1;
g. RTV (Rat theilovirus); h. Sendai virus; i. PVM; j. Mycoplasma
pulmonis; k. REO3; l. LCMV; m. CARB; n. Hantaan virus; o.
Clostridim piliforme; p. MAD1; q. MAD2; r. Encephalitozoom
cuniculi; and s. IDIR.
13. The method of claim 3, wherein the members of the population of
rodents are mice.
14. The method of claim 3, wherein the blood on the cards was
obtained from a facial vein.
15. The method of claim 3, wherein the members of the population of
rodents are rats.
16. The method of claim 3, wherein the identity of the at least one
of the rodent population and an individual member of the population
is associated with each of the plurality of blood collection
cards.
17. The method of claim 3, wherein the blood collection cards each
have at least one collection area having an absorbent material
suitable for holding about 10-40 .mu.L of whole blood.
18. The method of claim 3, wherein the blood collection cards each
have at least two collection areas.
19. The method of claim 18, wherein the at least two collection
areas are separated by a perforation that allows separation of the
cards such that a single sample area is on each separated card.
20. The method of claim 19, wherein the identity of at least one of
the rodent population and an individual member of the population is
associated with each sample area of each separated card.
21. The method of claim 13, wherein the step of analyzing the
extracted blood comprises analyzing for the presence or absence of
seven or more diseases selected from the group consisting of: a.
MHV; b. MVN (MMV); c. NS1 (Generic Parvovirus); d. MPV (MPV1-5); e.
MNV; f. TMEV; g. EDIM; h. Sendai virus; i. Mycoplasma pulmonis; j.
PVM; k. REO3; l. LCMV; m. Ectromelia virus; n. MAD1; o. MAD2; p.
Polyoma virus; q. Encephalitozoon cuniculi; r. CARB; s. Clostridium
piliforme; t. MCMV; u. K virus; v. Hantaan virus; w. Lactate
dehydrogenase-elevating virus; and x. MTV.
22. The method of claim 15, wherein the step of analyzing the
extracted blood comprises analyzing for the presence or absence of
seven or more diseases selected from the group consisting of: a.
RCV; b. NS1 (Generic Parvovirus); c. RPV; d. RMV; e. KRV; f. H-1;
g. RTV (Rat theilovirus); h. Sendai virus; i. PVM; j. Mycoplasma
pulmonis; k. REO3; l. LCMV; m. CARB; n. Hantaan virus; o.
Clostridim piliforme; p. MAD1; q. MAD2; r. Encephalitozoom
cuniculi; and s. IDIR.
Description
RELATED APPLICATIONS
[0001] This application is a continuation application of U.S.
patent application Ser. No. 14/538,381, filed Nov. 11, 2014, now
U.S. Pat. No. 8,945,945, which is a continuation of U.S. patent
application Ser. No. 14/089,103, filed Nov. 25, 2013, now U.S. Pat.
No. 8,927,298, which claims the benefit of U.S. Provisional
Application No. 61/776,560, filed Mar. 11, 2013.
FIELD
[0002] The invention relates to the collection and analysis of
biological samples from populations of animals. More particularly,
the invention is directed to the management of animal colonies
using dried blood samples from the members of the colony for
analysis of disease or phenotype of the colony.
BACKGROUND
[0003] Over the past five decades, great strides have been made in
the identification and eradication of infections from laboratory
rodents. As a result, most contemporary biomedical research rodent
colonies are relatively free of the pathogenic viruses, parasites,
bacteria, and fungi that cause clinical disease. However, some
microbes, especially those agents that cause subclinical disease,
remain in an enzootic state in many research colonies. These
agents, despite their insidious nature, have an impact on
physiologic parameters of the host and thus on the results of
animal experiments, independent of their pathogenic potential.
Therefore, timely and accurate diagnosis of infectious disease in
animal research models is critical to the success of biomedical
research. To this end, institutional veterinarians closely monitor
the health of research animals through periodic systematic
examination of sample groups of research animals against a
predetermined list of infectious agents. Rodent health monitoring
can generally be accomplished using a combination of molecular and
serological diagnostic assays. Molecular diagnostic tools provide a
real-time assessment of infection; whereas, serological tools
detect the presence of antibodies to infectious agents, thus,
providing an historical perspective of infectious disease exposure
over the life of the animal.
[0004] The current practice for collection of blood or serum for
serological evaluation of infectious disease in laboratory animals
includes: most commonly, euthanasia of animals for collection of at
least 100 .mu.L, of blood by cardiocentesis. Once collected the
whole blood sample is allowed to clot, which typically requires
2-12 hours, then whole blood is centrifuged and the serum is
separated from the cellular (clotted) fraction. Next, the serum is
shipped to a facility at refrigerated or frozen temperatures using
an overnight service (generally one or two pounds of ice packs are
required) in a STRYOFOAM.TM. shipping box.
[0005] This practice is inconvenient and expensive in light of the
amount of animal colonies and the number of analytes that must be
tested to ensure colony health and homogeneity. Accordingly, the
inventors have identified a need in the art to provide a simplified
and efficient method for sample collection and analysis to ensure
cost effective colony management.
SUMMARY
[0006] In one aspect, the disclosure is directed to a method for
managing a an animal colony. The method includes collecting blood
samples from a plurality of members of the colony on a plurality of
collection cards; allowing the blood samples to dry on the
collection cards; transporting the collection cards to a laboratory
as a single unit; extracting the samples from the cards; analyzing
the samples for the presence or absence of a biological marker; and
removing one or more of the members from the colony based upon the
presence or absence of the marker in the samples from the one or
more members. The biological marker may be a marker for an
infectious disease.
[0007] In various aspects of the disclosure each collection card in
the plurality of collection cards is labeled to identify the member
of the population associated with each sample on a card. The
samples may be analyzed in a multiplex immunoassay, for example,
and immunoassay that detects at least ten different analytes in the
samples. Each collection card may contain segments for collecting
up to, for example, five samples, fifty samples, or 100 samples.
The volume of blood of each sample collected on the card may be
about 10-40 .mu.L. Blood may be collected from the animal's lateral
saphenous vein, facial vein or temporal vein, and the blood may be
collected on the card without a collection device. The samples may
be dried and/or shipped at room temperature.
[0008] In a further aspect, the disclosure is directed to method of
determining the health status in a population of rodents. The
method includes providing a plurality of blood collection cards;
instructing the user to draw blood from an individual rodent;
instructing the user to apply the blood to one of the plurality of
blood collection cards; instructing the user to allow the blood
sample to dry on the collection card; instructing the user to
repeat the blood collection and drying at least once to provide a
plurality of blood collection cards spotted with blood from the
population of rodents; instructing the user to transport the
plurality of collection cards to a laboratory as a single unit;
extracting the samples from the cards; analyzing the samples for
the presence or absence of at least one biological marker for an
infectious disease; and reporting the results of the analysis back
to the user.
[0009] In various aspects, the members of the population of rodents
are mice, and the user is instructed to draw the blood from a
facial vein. In another aspect, the members of the population of
rodents are rats.
BRIEF DESCRIPTION OF THE FIGURES
[0010] FIGS. 1A and 1B shows an example of a prior art sample
collection card.
[0011] FIG. 2 shows an example of a sample collection card having
six spots for sample collection that can be separated (if desired)
by detachment along various perforations.
[0012] FIGS. 3A and 3B shows a top view of and a side view an
example of a sheet having a plurality of sample collection cards
that can be separated (if desired) by detachment along various
perforations.
DETAILED DESCRIPTION
[0013] The invention addresses the challenges associated with the
collection, identification and processing of voluminous numbers of
samples obtained from animal colonies, and in particular rodent
colonies. These colonies are maintained for research purposes and,
in many cases, it is critical that the members of the colonies have
particular phenotypes and health status. The testing of individual
members of the colonies for up 30 different bacterial and viral
agents and for desired phenotypic characteristics may be necessary
to ensure the health of the colony and its usefulness in testing,
for example, the efficacy of pharmaceutical agents on large
populations.
[0014] The use of the invention requires significantly less sample
from colony members than traditional sampling processes (.about.25
.mu.L vs. 100 .mu.L), making live (antemortem) sample collection
safer, more simple and more feasible (eliminates the need for
phlebotomists therefore enabling expanded or self-sampling).
Accordingly, in one aspect, the invention provides a library of
blood samples from a population of animals. The library includes
samples from a plurality of members of a colony. Typically, a
statically significant number of animals in the colony are tested
for particular phenotype or disease. For example, in very large
populations (e.g., up to 100,000 animals) as many to 400 animals
are tested (e.g., 2 animals per group of 50). In some populations,
all of the animals are tested. Collection of the test samples from
the individual members of the colony provides a library of samples
representative of the population.
[0015] In accordance with the invention, the library of samples is
collected on collection cards, which are typically absorbent and
inert fibrous thin sheet materials. In one particular embodiment,
the collection cards are WHATMAN.RTM. FTA.RTM. DMPK-C (GE
Healthcare Biosciences, Piscataway, N.J.), which have multiple
collection areas for samples of about 10-40 .mu.L. In other
embodiments, the spots can hold about 20-30 .mu.L, and in another
embodiment a 1 cm.sup.2 collection area holds approximately 25
.mu.L of whole blood. While the cards can be impregnated with
various chemicals (e.g., stabilizers, enzyme inhibitors, etc.), it
is preferred that the cards contain no impregnated chemicals. It
has also been found that use of the cards does not adversely
denature target proteins.
[0016] An example of a prior art blood collection card is shown in
FIGS. 1A and 1B. This card (Protein Saver.TM. 903.RTM. card by
Whatman) has five blood collection spots arranged on a continuous,
non-perforated web of material. In this embodiment, all blood
collection spots are present for purposes of collecting blood from
one patient/individual. After blood is spotted and preferably
dried, the top cover is tucked into the bottom panel as shown.
[0017] In rodent populations, blood is typically drawn from the
lateral saphenous vein, facial vein or the temporal vein. In one
aspect of the invention, collection is accomplished directly from
the vein on to the collection card without the use of a capillary
tube or other collection device by contacting the card directly
with the animal body at the site of the punctured vein or by
permitting a drop of blood to fall onto the desired area of the
card. Ideally, a single large drop should touch the card in order
to allow the sample to spread quickly and symmetrically on the
collection spot of the card surface to provide a reproducible,
uniform spot. Using the WHATMAN.RTM. FTA.RTM. DMPK-C cards, spot
formation is not essentially influenced by application speed or
direction, and provides minimal chromatographic separation in the
card. Spot area is generally proportional to sample volume, which
provides uniformity in sample size when the card is "punched" as a
first step of sample extraction from the card. In one embodiment,
the collection card is WHATMAN.RTM. FTA.RTM. 31 ETF PK paper.
[0018] For use with collection from the saphenous, facial or
temporal veins, it is desirable that each card contain only one
sample to avoid contamination between samples as the result of the
collection of blood from a live animal. When blood is collected
directed from the animal, it is spotted without anticoagulant.
Analytical labs, however, normally use blood containing EDTA or
another anticoagulant for controls and standards. For validated
assays it may be necessary to collect data showing the
anticoagulant to be unnecessary.
[0019] In another aspect, the invention provides that each
collection card identify the animal providing the sample. Animal
identification can be accomplished by several known means according
to animal colony and clinical laboratory management as generally
known in the art, including labels and barcodes containing
information that can be electronically stored and transmitted. In
one embodiment, a plurality of sample spots and room for subject
identifying information is provided on a sheet containing segments
that include the sample spot and identifying information.
Perforations between the spots allow for removal of segments to
provide individual collection cards for each subject containing the
sample and identification information. For instance, a sheet may
include 2 to 100, more particularly 2-10, for example 3, 4, 5, 6,
7, 8, or 9, segments that can be separated prior to or at the time
of sample collection.
[0020] Once the samples from the colony have been collected,
samples are dried, preferably in an environment with good air
circulation and low ambient humidity. Moderate heating may be
considered, but care should be taken so as not to damage the cards
or reduce analyte stability. Fans or vacuum desiccators may speed
the process, which generally takes about two hours at room
temperature. In another embodiment, the card can be immediately
folded to protect the sample without drying first, and without
smearing, disturbing or contaminating the sample.
[0021] Once the samples are dry, the cards can be arranged to be
transported, preferably gathered together as a single unit, for
providing information regarding one or more biological markers in
the population of animals. In this aspect, the library of samples
on the cards can be transported using commercially available
transportation and delivery services (e.g., U.S. Mail, FEDEX.RTM.,
UPS.RTM.) in standard delivery envelops without refrigeration to a
reference laboratory for analysis. Blood samples collected and
dried on the cards are generally stable for up to 7 days at room
temperature. Use of a desiccant in the shipping container can help
to avoid degradation. In some embodiments, library of collection
cards contains up 100, 200, 300 or 400 cards.
[0022] FIG. 2 shows a segmented collection card 6 having sample
spots. Card 10 has a sample panel 12 (depicted in gray), an
identification panel 14, a mid panel 16 and an end panel 18.
Optional fold scores are shown at 15. In one embodiment, the sample
panel 12 is WHATMAN.RTM. FTA.RTM. 31 ETF PK paper, while the other
panels can be a sturdy card stock such as 100 pound White Tag card
stock, for example about approximately 100 pound weight. In one
embodiment, the FTA.RTM. paper can be the length of the panel 12,
while the length of the card stock is that of the panels 14, 16 and
18, wherein the two sections are glued at line 17 where overlapping
at panel 12 (FTA.RTM. paper preferably on top of the card stock).
Card 10 is shown with six target sample circles 20A-F. Lateral
perforations 22 can be provided that run across the length of all
panels to allow the user to separate the card 10 such that a single
sample circle is on each separated card. As mentioned elsewhere in
this disclosure, the card can have multiple sample spots and
perforations. In use, after the sample(s) is applied, the first
panel 18, and then the panel 14, can be folded onto the panel 16 to
protect the sample(s) in 20A-F of the panel 12.
[0023] FIG. 3A shows an alternate embodiment of the present
invention wherein a sheet having multiple sample collection cards
that can be separated (if desired) by detachment along various
perforations. FIG. 3B depicts a cross sectional view of a sheet 100
having a base material 110 that is preferably a sturdy card
material (such as 100 pound White Tag), which is optionally
laminated to the collection material 122. A layer of a blood
collection material 122 is preferably glued on top of the base 110
as shown, and a top layer 111 (which is optionally laminated) of
the same material as the base material on top of the collection
material 122. As with FIG. 2, collection material 122 is any
material that can receive and secure a blood sample such as, for
example, FTA.RTM. paper. The top material 111 has a window 120
(e.g., 1.2 cm.times.2.0 cm), depicted in gray, which leaves the
collection material 122 exposed. The collection material 122 has
blood collection target spots 121 printed thereon to facilitate
application of the blood sample. The sheet 100 has a plurality of
horizontal perforations 114 and a plurality of vertical
perforations 112. By way of example only, if sheet 100 is 20
centimeters by 25 centimeters, then 55 individual cards (.about.5
centimeters by .about.1.8 centimeters) can be separated and used
for collection of samples from 55 individual subjects. Also shown
in FIG. 3A are optional fold lines 118 on sheet 100.
[0024] In use, the blood collection cards (each having a single
blood collection spot 20 or a spot 121 or more than one blood
collection spot) can be separated from card 10 or sheet 100.
Information about the blood donor can be applied to the card along
with the blood sample. After sample application, the card can be
optionally folded to protect the sample and transported to a
testing facility. Results of tests run on the blood sample can be
matched with the donor and the information transmitted back to the
facility (e.g., via e-mail, computer network, internet, in writing,
etc.) where the blood sample was taken. In the case of rodent
colony testing, one or more tests that show positive for the
presence of a non-desirable condition (e.g., infection, see below),
may result in the segregation, quarantine and/or euthanasia of
rodents in the colony.
[0025] The arrangement of the cards does not necessarily require
spacers between the cards to avoid contact between dried blood
samples. However, it may be desirable to include a flap that can be
folded over once or twice so that the sample spot is covered on at
least one side of the spot. The flap can ensure that sample spots
on different cards are not in direct contact during shipping. As
shown in FIG. 2, the cards can contain one or more fold scores or
other indications of where the card should be folded to ensure
isolation of the spot and, in some instances, allow access to
identifying information without unfolding the flap.
[0026] At the clinical laboratory, samples are removed from the
cards by punching the cards with a punch that provides a core
containing a uniform sample size when the sample has been properly
collected and the size of the punch is smaller than the sample
spots. Typical punch sizes for samples of up to 20 microliters are
3-10 mm. Larger samples representing up to 100 microliters of
properly collected blood can be obtained with a larger punch, for
example 9 mm. If capturing total sample volume is desirable over
uniformity of sample size, a punch larger than the spots, or
scissors, can be used to ensure that the core contains the entire
sample volume.
[0027] Once the sample has been punched from the cards, the samples
can be extracted from the core using known solvents. For small
molecules, the solvent can be anything that is a solvent for the
analyte. Methanol and acetonitrile are widely used, either straight
or mixed with water. Water itself may also be used for extremely
polar analytes. Extraction of ionizable analytes is often improved
by pH adjustment--increasing the charge to improve solubility in
water or reducing it to promote solubility in organic solvents. In
some instances, extraction of even moderately polar analytes is
increased by adding perhaps 10-15% water to methanol, sometimes by
adding water to the dry spot first, then allowing to soak a few
minutes before adding organic solvent. An extremely hydrophobic
analyte may be best extracted with a nonpolar solvent such as
hexane, also providing some cleanup by leaving polar contaminants
undissolved in the punch.
[0028] For peptides and proteins, aqueous buffers with pH and salt
concentration to promote protein stability can be used. The
addition of a non-ionic detergent, such as 0.1% TWEEN.TM.-20 or
TRITON.TM. X-100 detergents may be desirable. Removal may require
incubation with gentle mixing for one to several hours. Hydrophobic
peptides will probably extract better with some methanol or
acetonitrile added.
[0029] In on embodiment, a 9 mm punch is removed, using an
appropriate 9 mm single hole punch, from the card, placed in a
sterile tube. Antibodies are eluted with 100 microliters of buffer
containing Tris-buffered saline with 1 mM EDTA. The tube is placed
in the refrigerator overnight to allow efficient antibody elution
from the membrane.
[0030] Once the library samples are extracted, the samples are
analyzed for the presence or absence of a biological marker. For
example, mouse colonies can be tested for the following infectious
agents, and the samples may be tested in various subsets (panels)
as exemplified in Table 1.
TABLE-US-00001 TABLE 1 Mouse Panel Test A B C D E F MHV X X X X X X
MVM (MMV) X X X X X X NS1 (Generic Parvovirus) X X X X X X MPV
(MPV1-5) X X X X X X MNV X X X X X X TMEV X X X X X X EDIM X X X X
X X Sendai virus X X X X X Mycoplasma pulmonis X X X X X PVM X X X
X REO3 X X X X LCMV X X X X Ectromelia virus X X X X MAD1 X X X
MAD2 X X X Polyoma virus X X X Encephalitozoon cuniculi X X CARB X
X Clostridium piliforme X X MCMV X X K virus X Hantaan virus X
Lactate dehydrogenase-elevating virus X MTV(IFA) X
[0031] Similarly, for rat colonies, the analytes and panels are
exemplified in Table 2.
TABLE-US-00002 TABLE 2 Rodent Panel Test A B C D E RCV X X X X X
NS1 (Generic Parvovirus) X X X X X RPV X X X X X RMV X X X X X KRV
X X X X X H-1 X X X X X RTV (Rat theilovirus) X X X X X Sendai
virus X X X X PVM X X X X Mycoplasma pulmonis X X X X REO3 X X X
LCMV X X X CARB X X Hantaan virus X X Clostridium piliforme X X
MAD1 X X MAD2 X Encephalitozoon cuniculi X IDIR X
[0032] Animal colonies of other species can be analyzed for panels
of markers appropriate for the species.
[0033] Overall sensitivity for small molecules is very much a
function of analyte, matrix interferences, chromatography
conditions and mass spectrometer capabilities. In general, values
in the range of 0.1-10 ng/mL can be obtained from single 3 mm
punches. Proteins can be extracted from blood spots and detected by
immunoassay with sensitivity comparable to standard plasma or serum
samples.
[0034] In one aspect, the extracted samples are analyzed in a
Multiplex Fluorescent Immunoassay (MFI) that is based both on
bead-based immunoassay and flow cytometry. Purified antigen or
control preparations are covalently linked to one of, for example,
100 different types of polystyrene beads, which vary slightly in
the intensity of their color. If IgG antibody to a particular
antigen is present, then it will bind to the antigen on a specific
bead and will then be detected by subsequent binding of goat
anti-species antibody conjugated to a fluorochrome (e.g.,
R-phycoerythrin). The reader channels single beads through a dual
laser detector which simultaneously determines both the bead type
by the internal dye combination and the fluorescent intensity
associated with each individual bead. The fluorescent intensity
associated with each of the individual beads of each type are used
in the determination of each MFI value. Side-by-side testing of
thousands of individual results from hundreds of samples show
overall correlation between MFI and ELISA is greater than 99.5% for
both mouse and rat samples. In general, MFI is more sensitive than
ELISA and is less prone to false positive results. MFI requires
only 1.0 .mu.L of undiluted serum (5.0 .mu.L of 1:5 diluted serum)
regardless of the number of tests requested.
[0035] The ability to use small sample sizes for testing several
analytes in a sample using MFI sample coupled with the invention
including library of sample collection cards allows for the
comprehensive and convenient analysis of a colony of survival-bled
(antemortem) animals. The results of the colony analysis can be
transmitted directed to the colony manager by electronic
communications, including e-mail and smart phone applications, so
that that laboratory manager has immediate access to data regarding
the colony or individual room of a colony.
[0036] In other embodiments, the eluant is then evaluated by other
know immunoassay techniques known to those of skill in the art
(e.g., IFA and western blot).
[0037] In one aspect, the invention is directed to a method of
managing a rodent colony. The method analysis of biological markers
for disease or phenotype within the colony using sample collection
cards, and sample collection and analysis as described herein.
Colony management may include removing members from the colony that
test positive or negative for the biological marker.
[0038] The following are provided for exemplification purposes only
and are not intended to limit the scope of the invention described
in broad terms above. All references cited in this disclosure are
incorporated herein by reference.
EXAMPLES
Example 1
Monitoring a Rodent Colony for Infectious Agents
[0039] Routine rodent health monitoring for infectious agent
exposure is accomplished by serologically evaluating rodent serum
samples for the presence of antibodies formed as part of the immune
response to infection. To accomplish this, blood is collected, via
venipuncture, from sample groups of research animals (e.g.,
.about.10% of the rodent colony of interest) and spotted onto
membrane cards labeled with unique animal identification codes to
allow for later identification of the animal. Once the blood
samples from the rodent research colony have been collected and
samples are dried, the cards are transported, using commercially
available transportation and delivery services (e.g., U.S. Mail,
FEDEX.RTM. or UPS.RTM.) in standard delivery envelops without
refrigeration, to a reference laboratory for analysis. Samples are
tested against a predetermined list of indicators (see for example
Table 1 and Table 2) of an infection and results are reported to
the submitter. When an infectious disease outbreak is detected, the
infected animals are identified using the unique identification
codes and quarantined. Additional steps may be taken to ascertain
the extent of the outbreak and to eliminate/control the infectious
agent. Ultimately, it may be necessary to re-derive or restock the
colony with disease-free animals.
[0040] Although various specific embodiments of the present
invention have been described herein, it is to be understood that
the invention is not limited to those precise embodiments and that
various changes or modifications can be affected therein by one
skilled in the art without departing from the scope and spirit of
the invention. The examples given above are merely illustrative and
are not meant to be an exhaustive list of all possible embodiments,
applications or modifications of the invention. Thus, various
modifications and variations of the described invention will be
apparent to those skilled in the art without departing from the
scope and spirit of the invention. Although the invention has been
described in connection with specific embodiments, it should be
understood that the invention as claimed should not be unduly
limited to such specific embodiments. Indeed, various modifications
of the described modes for carrying out the invention which are
obvious to those skilled in animal colony management, molecular
biology, immunology, chemistry, biochemistry or in the relevant
fields are intended to be within the scope of the appended
claims.
[0041] Any numerical values recited herein include all values from
the lower value to the upper value in increments of one unit
provided that there is a separation of at least two units between
any lower value and any higher value. As an example, if it is
stated that the concentration of a component or value of a process
variable such as, for example, size, angle size, pressure, time and
the like, is, for example, from 1 to 90, specifically from 20 to
80, more specifically from 30 to 70, it is intended that values
such as 15 to 85, 22 to 68, 43 to 51, 30 to 32, etc. are expressly
enumerated in this specification. For values that are less than
one, one unit is considered to be 0.0001, 0.001, 0.01 or 0.1 as
appropriate. These are only examples of what is specifically
intended and all possible combinations of numerical values between
the lowest value and the highest value enumerated are to be
considered to be expressly stated in this application in a similar
manner.
[0042] The disclosures of all references and publications cited
herein are expressly incorporated by reference in their entireties
to the same extent as if each were incorporated by reference
individually.
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