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Chapter 35 : Multiplex Cytokine Assays

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Multiplex Cytokine Assays, Page 1 of 2

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Abstract:

Cytokines are low-molecular-weight proteins representing important components of inflammatory and immune reactions (1). In response to multiple stimuli (2), cytokines are rapidly induced and secreted into the extracellular milieu. However, in some situations, cytokines are constitutively present. Cytokines exert numerous biological activities which are critical for host defense, physiologic responses to stress, and immune surveillance. Cytokines, along with complement, are considered to be part of the innate immune system. The world of cytokine biology has exploded in the past decades. It can be said without hyperbole that cytokines are critical from birth (gestation) (3) to death (apoptosis) (4).

Citation: Duffy E, Remick D. 2016. Multiplex Cytokine Assays, p 324-337. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch35
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Image of FIGURE 1
FIGURE 1

Direct ELISA. In the direct ELISA, the analyte is first bound to the bottom of the microtiter well. In this example, the analyte is IL-6. Unbound IL-6 is washed away, and excess protein binding sites are blocked in order to reduce background. In the next step, biotin-labeled antibody directed against IL-6 is added; the small circle represents the biotin moiety directly attached to the antibody. After washing, streptavidin (SA) conjugated to horseradish peroxidase (HRP) is added and a final wash is performed. Following addition of a colorimetric substrate, the color develops. The intensity of the color development is directly proportional to the amount of IL-6 in the first step.

Citation: Duffy E, Remick D. 2016. Multiplex Cytokine Assays, p 324-337. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch35
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Image of FIGURE 2
FIGURE 2

Indirect or sandwich ELISA. This ELISA shares many features with the assay described in the legend to Fig. 1 . The major difference is in the first step, in which an antibody (Ab) directed against the analyte is bound to the bottom of the microtiter well. This antibody is termed the capture or coating antibody. Following addition of the analyte, the biotin-labeled detection antibody (biotin-Ab) is applied, followed by horseradish peroxidase (HRP)-conjugated streptavidin (SA-HRP). Color development proceeds, and the intensity of the color is directly related to the amount of the analyte. This is termed a sandwich ELISA because the analyte is sandwiched between two antibodies.

Citation: Duffy E, Remick D. 2016. Multiplex Cytokine Assays, p 324-337. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch35
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Image of FIGURE 3
FIGURE 3

Sequential ELISA. A sample containing a mixture of cytokines is applied to an IL-6 ELISA. IL-6 binds to the anti-IL-6 antibody attached to the bottom of the well. When the sample is removed, cytokines which are not IL-6, such as tumor necrosis factor (TNF), may be used in a subsequent ELISA. Ab, antibody; B, biotin moiety; biotin-Ab, biotin-labeled antibody; HRP, horseradish peroxidase; SA-HRP, horseradish peroxidase-conjugated streptavidin.

Citation: Duffy E, Remick D. 2016. Multiplex Cytokine Assays, p 324-337. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch35
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Image of FIGURE 4
FIGURE 4

Example of a microarray. Panel A shows six individual nitrocellulose pads arranged on a glass slide. Each pad has been spotted in an 8-by-12 format. The black area between the pads is the place where a silicon gasket was adhered to the surface to allow each of the wells to function individually, similar to the way in which an individual well in a 96-well plate is independent from its neighbors. The antibodies have been spotted in an identical manner onto each of the nitrocellulose pads. Panel B is an enlarged picture of one of the individual pads and highlights the detail of the spots. The antibodies have been spotted in quadruplicate. Each individual spot has a diameter of 150 μm, and the distance between the spots is 300 μm. The total volume delivered to each spot was 350 to 367 pl. In the far right column of spots, those in the top eight positions are extremely bright and the lower four have virtually no signal. This line of eight plus four spots may be used for alignment of the image from the protein chip. The intensity of the individual spots may be quantified and used to determine the cytokine concentration in the sample. This image shows excellent reproducibility of the quadruple spots.

Citation: Duffy E, Remick D. 2016. Multiplex Cytokine Assays, p 324-337. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch35
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Image of FIGURE 5
FIGURE 5

Specificity of the microarray. For this microarray, 18 different cytokines were tested. A cocktail containing recombinant cytokines was prepared, but IL-12 was not added to the cocktail. There is a strong signal from all the other cytokines, but no fluorescence was observed on the array for IL-12. This demonstrates the specificity for the array.

Citation: Duffy E, Remick D. 2016. Multiplex Cytokine Assays, p 324-337. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch35
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Image of FIGURE 6
FIGURE 6

Mathematical modeling of the standard curve. Data from a microarray standard curve were used to generate a mathematical model for calculations for unknown samples. In panel A, the number of relative light units (RLU) and the concentration of the cytokine (IL-8) were plotted on a linear-linear scale. The correlation coefficient () was not very precise. In panel B, the concentration of the IL-8 was plotted on a log scale and the number of RLU was plotted on a linear scale. This resulted in even worse correlation. In panel C, both the number of RLU and the IL-8 concentration were plotted on a log scale and a very good linear regression could be fitted. However, the optimal modeling of the curve was obtained when the number of RLU was plotted on a linear scale, the IL-8 concentration was plotted on a log scale, and a fourth-order polynomial regression line was used. The results for all four panels were obtained using the same data from the IL-8 standard curve, but similar results are observed with most other cytokines.

Citation: Duffy E, Remick D. 2016. Multiplex Cytokine Assays, p 324-337. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch35
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Image of FIGURE 7
FIGURE 7

Increase in signal intensity with smaller spot size. On the left, each black dot represents the analyte bound to an antibody, which spreads across the entire well. On the right, the same number of antibodies are bound to the analyte, but they now occupy a smaller space, with a resulting increase in signal intensity.

Citation: Duffy E, Remick D. 2016. Multiplex Cytokine Assays, p 324-337. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch35
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Image of FIGURE 8
FIGURE 8

Schematic representation of the principle of a flow cytometric bead-based assay. The capture antibody specific for the analyte, in this example IL-6, is attached to a bead. The IL-6 binds to this antibody, which is then followed by a detection antibody which is biotinylated. Streptavidin conjugated to phycoerythrin is then added, and the entire complex is analyzed with the flow cytometer.

Citation: Duffy E, Remick D. 2016. Multiplex Cytokine Assays, p 324-337. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch35
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Image of FIGURE 9
FIGURE 9

Example of the readout from a bead-based flow cytometry assay. In this example, the individual beads have different concentrations of two fluorescent dyes. The groups of beads are detected on the basis of the fluorescence intensities. The individual dots represent individual beads, and the darker-colored dots are doublets. The clear areas around collections of beads indicate those areas used for analysis. In this example, 22 individual cytokines have been detected. The intensity of the fluorescence of each individual bead is captured in a third fluorescent channel and compared to the standard curve in order to determine the concentration of the cytokine.

Citation: Duffy E, Remick D. 2016. Multiplex Cytokine Assays, p 324-337. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch35
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Image of FIGURE 10
FIGURE 10

Example of a standard curve generated from the bead-based flow cytometry assay. In this example, a standard curve using recombinant mouse IL-1β is displayed. MFI, mean fluorescence intensity.

Citation: Duffy E, Remick D. 2016. Multiplex Cytokine Assays, p 324-337. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch35
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References

/content/book/10.1128/9781555818722.ch35
1. Remick DG, Friedland JS. 1997. Cytokines in Health and Disease, 2nd ed. Marcel Dekker, Inc, New York, NY.
2. Jin C, Flavell RA. 2013. Innate sensors of pathogen and stress: linking inflammation to obesity. J Allergy Clin Immunol 132:287294.[CrossRef].[PubMed]
3. Chaouat G, Ledee-Bataille N, Dubanchet S, Zourbas S, Sandra O, Martal J. 2004. TH1/TH2 paradigm in pregnancy: paradigm lost? Cytokines in pregnancy/early abortion: reexamining the TH1/TH2 paradigm. Int Arch Allergy Immunol 134:93119.[CrossRef].[PubMed]
4. Martinon F, Tschopp J. 2004. Inflammatory caspases: linking an intracellular innate immune system to autoinflammatory diseases. Cell 117:561574.[CrossRef].[PubMed]
5. Mease PJ, Fleischmann R, Deodhar AA, Wollenhaupt J, Khraishi M, Kielar D, Woltering F, Stach C, Hoepken B, Arledge T, van der Heijde D. 2014. Effect of certolizumab pegol on signs and symptoms in patients with psoriatic arthritis: 24-week results of a Phase 3 double-blind randomised placebo-controlled study (RAPID-PsA). Ann Rheum Dis 73:4855.[CrossRef].[PubMed]
6. Akalin H, Akdis AC, Mistik R, Helvaci S, Kilicturgay K. 1994. Cerebrospinal fluid interleukin-1 beta/interleukin-1 receptor antagonist balance and tumor necrosis factor-alpha concentrations in tuberculous, viral and acute bacterial meningitis. Scand J Infect Dis 26:667674.[PubMed].[CrossRef]
7. Atwell DM, Grichnik KP, Newman MF, Reves JG, McBride WT. 1998. Balance of proinflammatory and antiinflammatory cytokines at thoracic cancer operation. Ann Thorac Surg 66:11451150.[PubMed].[CrossRef]
8. Khan SS, Smith MS, Reda D, Suffredini AF, McCoy JP Jr. 2004. Multiplex bead array assays for detection of soluble cytokines: comparisons of sensitivity and quantitative values among kits from multiple manufacturers. Cytometry B Clin Cytom 61:3539.[CrossRef].[PubMed]
9. Kohler K, Seitz H. 2012. Validation processes of protein biomarkers in serum—a cross platform comparison. Sensors (Basel) 12:1271012728.[CrossRef].[PubMed]
10. Brody EN, Willis MC, Smith JD, Jayasena S, Zichi D, Gold L. 1999. The use of aptamers in large arrays for molecular diagnostics. Mol Diagn 4:381388.[CrossRef].[PubMed]
11. Zhu H, Snyder M. 2003. Protein chip technology. Curr Opin Chem Biol 7:5563.[PubMed].[CrossRef]
12. Yalow RS, Berson SA. 1959. Assay of plasma insulin in human subjects by immunological methods. Nature 184(Suppl 21):16481649.[PubMed].[CrossRef]
13. Nielsen UB, Geierstanger BH. 2004. Multiplexed sandwich assays in microarray format. J Immunol Methods 290:107120.[CrossRef].[PubMed]
14. Vignali DA. 2000. Multiplexed particle-based flow cytometric assays. J Immunol Methods 243:243255.[PubMed].[CrossRef]
15. Herzyk DJ, Berger AE, Allen JN, Wewers MD. 1992. Sandwich ELISA formats designed to detect 17 kDa IL-1 beta significantly underestimate 35 kDa IL-1 beta. J Immunol Methods 148:243254.[PubMed].[CrossRef]
16. Herzyk DJ, Wewers MD. 1993. ELISA detection of IL-1 beta in human sera needs independent confirmation. False positives in hospitalized patients. Am Rev Respir Dis 147:139142.[CrossRef].[PubMed]
17. Nemzek JA, Siddiqui J, Remick DG. 2001. Development and optimization of cytokine ELISAs using commercial antibody pairs. J Immunol Methods 255:149157.[PubMed].[CrossRef]
18. Lewkowich IP, Campbell JD, HayGlass KT. 2001. Comparison of chemiluminescent assays and colorimetric ELISAs for quantification of murine IL-12, human IL-4 and murine IL-4: chemiluminescent substrates provide markedly enhanced sensitivity. J Immunol Methods 247:111118.[PubMed].[CrossRef]
19. Obenauer-Kutner LJ, Jacobs SJ, Kolz K, Tobias LM, Bordens RW. 1997. A highly sensitive electrochemiluminescence immunoassay for interferon alfa-2b in human serum. J Immunol Methods 206:2533.[PubMed].[CrossRef]
20. Petrovas C, Daskas SM, Lianidou ES. 1999. Determination of tumor necrosis factor-alpha (TNF-alpha) in serum by a highly sensitive enzyme amplified lanthanide luminescence immunoassay. Clin Biochem 32:241247.[PubMed].[CrossRef]
21. Rongen HA, van der Horst HM, van Oosterhout AJ, Bult A, van Bennekom WP. 1996. Application of xanthine oxidase-catalyzed luminol chemiluminescence in a mouse interleukin-5 immunoassay. J Immunol Methods 197:161169.[PubMed].[CrossRef]
22. Siddiqui J, Remick DG. 2003. Improved sensitivity of colorimetric compared to chemiluminescence ELISAs for cytokine assays. J Immunoassay Immunochem 24:273283.[CrossRef].[PubMed]
23. Chiswick EL, Duffy E, Japp B, Remick D. 2012. Detection and quantification of cytokines and other biomarkers. Methods Mol Biol 844:1530.[CrossRef].[PubMed]
24. O'Connor KA, Holguin A, Hansen MK, Maier SF, Watkins LR. 2004. A method for measuring multiple cytokines from small samples. Brain Behav Immun 18:274280.[CrossRef]
25. Osuchowski MF, Remick DG. 2006. The repetitive use of samples to measure multiple cytokines: the sequential ELISA. Methods 38:304311.[CrossRef].[PubMed]
26. Osuchowski MF, Siddiqui J, Copeland S, Remick DG. 2005. Sequential ELISA to profile multiple cytokines from small volumes. J Immunol Methods 302:172181.[CrossRef].[PubMed]
27. Olson JA Jr. 2004. Application of microarray profiling to clinical trials in cancer. Surgery 136:519523.[CrossRef].[PubMed]
28. Tonkinson JL, Stillman BA. 2002. Nitrocellulose: a tried and true polymer finds utility as a post-genomic substrate. Front Biosci 7:c1c12.[PubMed]
29. Remick DG, Bolgos GR, Siddiqui J, Shin J, Nemzek JA. 2002. Six at six: interleukin-6 measured 6 h after the initiation of sepsis predicts mortality over 3 days. Shock 17:463467.[PubMed].[CrossRef]
30. Turnbull IR, Javadi P, Buchman TG, Hotchkiss RS, Karl IE, Coopersmith CM. 2004. Antibiotics improve survival in sepsis independent of injury severity but do not change mortality in mice with markedly elevated interleukin 6 levels. Shock 21:121125.[CrossRef].[PubMed]
31. Natarajan S, Remick DG. 2008. The ELISA Standard Save: calculation of sample concentrations in assays with a failed standard curve. J Immunol Methods 336:242245.[CrossRef].[PubMed]
32. Natarajan S, Remick DG. 2013. ELISA rescue protocol: recovery of sample concentrations from an assay with an unsuccessful standard curve. Methods 61:6972.[CrossRef].[PubMed]
33. Schweitzer B, Roberts S, Grimwade B, Shao W, Wang M, Fu Q, Shu Q, Laroche I, Zhou T, Tchernev VT, Christiansen J, Valleca M, Kingsmore SF. 2002. Multiplexed protein profiling on microarrays by rolling-circle amplification. Nature Biotechnol 20:359365.[CrossRef]
34. Sreekumar A, Nyati MK, Varambally S, Barrette TR, Ghosh D, Lawrence TS, Chinnaiyan AM. 2001. Profiling of cancer cells using protein microarrays: discovery of novel radiation-regulated proteins. Cancer Res 61:75857593.[PubMed]
35. Mendoza LG, McQuary P, Mongan A, Gangadharan R, Brignac S, Eggers M. 1999. High-throughput microarray-based enzyme-linked immunosorbent assay (ELISA). Biotechniques 27:778780, 782–776, 788.[PubMed]
36. Wiese R, Belosludtsev Y, Powdrill T, Thompson P, Hogan M. 2001. Simultaneous multianalyte ELISA performed on a microarray platform. Clin Chem 47:14511457.[PubMed]
37. Moody MD, Van Arsdell SW, Murphy KP, Orencole SF, Burns C. 2001. Array-based ELISAs for high-throughput analysis of human cytokines. Biotechniques 31:186190, 192184.[PubMed]
38. Huang RP. 2003. Protein arrays, an excellent tool in biomedical research. Front Biosci 8:d559d576.[PubMed].[CrossRef]
39. Roda A, Guardigli M, Russo C, Pasini P, Baraldini M. 2000. Protein microdeposition using a conventional ink-jet printer. Biotechniques 28:492496.[PubMed]
40. Huang RP. 2001. Detection of multiple proteins in an antibody-based protein microarray system. J Immunol Methods 255:113.[PubMed].[CrossRef]
41. Huang RP. 2001. Simultaneous detection of multiple proteins with an array-based enzyme-linked immunosorbent assay (ELISA) and enhanced chemiluminescence (ECL). Clin Chem Lab Med 39:209214.[CrossRef].[PubMed]
42. Huang RP, Huang R, Fan Y, Lin Y. 2001. Simultaneous detection of multiple cytokines from conditioned media and patient's sera by an antibody-based protein array system. Anal Biochem 294:5562.[CrossRef].[PubMed]
43. Lin Y, Huang R, Santanam N, Liu YG, Parthasarathy S, Huang RP. 2002. Profiling of human cytokines in healthy individuals with vitamin E supplementation by antibody array. Cancer Lett 187:1724.[PubMed].[CrossRef]
44. Copeland S, Siddiqui J, Remick D. 2004. Direct comparison of traditional ELISAs and membrane protein arrays for detection and quantification of human cytokines. J Immunol Methods 284:99106.[PubMed].[CrossRef]
45. Phillips TM. 2004. Rapid analysis of inflammatory cytokines in cerebrospinal fluid using chip-based immunoaffinity electrophoresis. Electrophoresis 25:16521659.[CrossRef].[PubMed]
46. Dossus L, Becker S, Achaintre D, Kaaks R, Rinaldi S. 2009. Validity of multiplex-based assays for cytokine measurements in serum and plasma from “non-diseased” subjects: comparison with ELISA. J Immunol Methods 350:125132.[CrossRef].[PubMed]
47. Frengen J, Schmid R, Kierulf B, Nustad K, Paus E, Berge A, Lindmo T. 1993. Homogeneous immunofluorometric assays of alpha-fetoprotein with macroporous, monosized particles and flow cytometry. Clin Chem 39:21742181.[PubMed]
48. McHugh TM. 1994. Flow microsphere immunoassay for the quantitative and simultaneous detection of multiple soluble analytes. Methods Cell Biol 42:575595.[PubMed].[CrossRef]
49. Iannone MA, Taylor JD, Chen J, Li MS, Rivers P, Slentz-Kesler KA, Weiner MP. 2000. Multiplexed single nucleotide polymorphism genotyping by oligonucleotide ligation and flow cytometry. Cytometry 39:131140.[PubMed].[CrossRef]
50. Iannone MA, Taylor JD, Chen J, Li MS, Ye F, Weiner MP. 2003. Microsphere-based single nucleotide polymorphism genotyping. Methods Mol Biol 226:123134.[CrossRef].[PubMed]
51. Pei R, Lee J, Chen T, Rojo S, Terasaki PI. 1999. Flow cytometric detection of HLA antibodies using a spectrum of microbeads. Hum Immunol 60:12931302.[PubMed].[CrossRef]
52. Kellar KL, Douglass JP. 2003. Multiplexed microsphere-based flow cytometric immunoassays for human cytokines. J Immunol Methods 279:277285.[PubMed].[CrossRef]
53. Morgan E, Varro R, Sepulveda H, Ember JA, Apgar J, Wilson J, Lowe L, Chen R, Shivraj L, Agadir A, Campos R, Ernst D, Gaur A. 2004. Cytometric bead array: a multiplexed assay platform with applications in various areas of biology. Clin Immunol 110:252266.[CrossRef].[PubMed]
54. Swartzman EE, Miraglia SJ, Mellentin-Michelotti J, Evangelista L, Yuan PM. 1999. A homogeneous and multiplexed immunoassay for high-throughput screening using fluorometric microvolume assay technology. Anal Biochem 271:143151.[CrossRef].[PubMed]
55. Prabhakar U, Eirikis E, Davis HM. 2002. Simultaneous quantification of proinflammatory cytokines in human plasma using the LabMAP assay. J Immunol Methods 260:207218.[PubMed].[CrossRef]
56. Prabhakar U, Eirikis E, Reddy M, Silvestro E, Spitz S, Pendley C II, Davis HM, Miller BE. 2004. Validation and comparative analysis of a multiplexed assay for the simultaneous quantitative measurement of Th1/Th2 cytokines in human serum and human peripheral blood mononuclear cell culture supernatants. J Immunol Methods 291:2738.[CrossRef].[PubMed]
57. Churchman SM, Geiler J, Parmar R, Horner EA, Church LD, Emery P, Buch MH, McDermott MF, Ponchel F. 2012. Multiplexing immunoassays for cytokine detection in the serum of patients with rheumatoid arthritis: lack of sensitivity and interference by rheumatoid factor. Clin Exp Rheumatol 30:534542.[PubMed]
58. Weixelbaumer KM, Raeven P, Redl H, van Griensven M, Bahrami S, Osuchowski MF. 2010. Repetitive low-volume blood sampling method as a feasible monitoring tool in a mouse model of sepsis. Shock 34:420426.[CrossRef].[PubMed]
59. Saiki RK, Gelfand DH, Stoffel S, Scharf SJ, Higuchi R, Horn GT, Mullis KB, Erlich HA. 1988. Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science 239:487491.[PubMed].[CrossRef]
60. Chen Y, Dong L, Weng D, Liu F, Song L, Li C, Tang W, Chen J. 2013. 1,3-β-glucan affects the balance of Th1/Th2 cytokines by promoting secretion of anti-inflammatory cytokines in vitro. Mol Med Rep 8:708712.[CrossRef].[PubMed]
61. Pusterla N, Madigan JE, Leutenegger CM. 2006. Real-time polymerase chain reaction: a novel molecular diagnostic tool for equine infectious diseases. J Vet Intern Med 20:312.[PubMed].[CrossRef]

Tables

Generic image for table
TABLE 1

Cost analysis for cytokine measurements

Citation: Duffy E, Remick D. 2016. Multiplex Cytokine Assays, p 324-337. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch35
Generic image for table
TABLE 2

Sliding-scale cost of measuring cytokines

Citation: Duffy E, Remick D. 2016. Multiplex Cytokine Assays, p 324-337. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch35
Generic image for table
TABLE 3

Sample volumes required for performing assays

Citation: Duffy E, Remick D. 2016. Multiplex Cytokine Assays, p 324-337. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch35

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