1.
【GeniePlex多重免疫檢測技術】 GeniePlex Multiplex Assays-Assaygenie
GeniePlex多重免疫檢測技術 (Multiplex ELISA Array, Assay Genie Multiplex ELISA)
GeniePlex 多重檢測是一種基於微珠的多重免疫檢測技術，幾乎可以在任何流式細胞儀上同時定量檢測單個 15ul 樣本中的多達 24 種分析物。可以一個Well同時偵測24 種不同的 ELISA！目前產品種類很多，包含:apoptosis, inflammation, angiogenesis, immune response 和migration, 可由人類、小鼠、大鼠、非人類靈長類動物和犬科動物的 400 多種分析物中進行選擇。大多數檢測的靈敏度低於 10.0 pg/ml。測定時間約 4個小時。測定內 CV：<10%，測定間 <20% ( Intra-Assay CV: <10%, Inter-Assay < 20%) 。
已經開發了幾種基於抗體的測定平台(antibody-based assay platforms)作為 ELISA 的替代品，用於同時定量分析單個樣品中的分析物，也稱為多重測定(multiplex assays)。最常見的多重免疫(multiplex immunoassays)分析平台使用基於流動的技術(flow-based technology)和抗體包被的珠子(antibody-coated beads)。珠子可用於同時測量單個樣品中的多種分析物(multiple analytes)。
在多重分析(multiplex assay)中，特定分析物的特異性抗體與分析中的一組珠子相連。然後添加另一種抗體，該抗體附有螢光報告基因。每組珠子都有一種相關的顏色，從而可以同時檢測單個樣品中的多種分析物。分析物的檢測可以通過流式細胞儀讀取，因為珠子可以通過各種螢光特徵檢測。可以根據檢測到的不同珠子顏色的數量來量化分析物的量。
Assay Genie Multiplex ELISA 試劑盒允許研究人員通過流式細胞術使用低至 15μL 的量對多達 24 種分析物進行多重分析！
多重ELISA原理-Assay Genie Multiplex ELISA 利用按大小和不同螢光強度水平區分的多個珠群。多重尺寸的珠子和每個珠子尺寸的多個螢光強度水平，多重技術可以在單個反應中同時測量多達 24 種分析物。多重 ELISA 只需要 15μL 的樣品。反應中的珠子數量由配備單個 488nm 激光器或雙 488nm 和 633/640nm 激光器的流式細胞儀確定。珠分類染料的最大發射波長為 700 nm。
基於珠子的免疫測定類似於夾心 ELISA 的原理，每個珠子群與捕獲樣品中感興趣的蛋白質（例如細胞因子）的特異性捕獲抗體偶聯。通過針對蛋白質二級表位的生物素化抗體檢測捕獲的分析物的量，然後進行鏈黴親和素-R-藻紅蛋白處理。珠子上的 R-藻紅蛋白的螢光強度在流式細胞儀上進行量化。通過將螢光信號與由已知濃度的分析物的連續稀釋產生的標準曲線的螢光信號進行比較，可以獲得樣品中感興趣的蛋白質的濃度。測定程序包括 60 分鐘的抗原和捕獲抗體偶聯珠孵育步驟、30 分鐘的生物素化檢測孵育步驟和 20 分鐘的鏈黴親和素 PE 孵育步驟。
樣品形式:
Cell Culture Supernatants、Plasma、Serum、Tissue Lysates、 Cell Lysates
Features & Benefits
- Analyze: Multiplex 2-24 analytes in a single sample!
- Fast: 2 hour protocol!
- Sensitive: Measure as low as <10 pg/ml of each analyte
- Dynamic: Lower limit < 20 pg/mL | Upper limit > 5,000 pg/mL
- High Precision Intra-assay CV: < 10% | Inter-assay CV: < 20%
- Low Volumes: Use as little as 15μl of sample
- Validated: All assays fully tested for cross-reactivity in our lab
Multiplex vs. Luminex?
The Assay Genie Multiplex ELISA gives excellent performance versus Luminex technology. The Human TH1/TH2/TH17 7-Plex ELISA and a Luminex multiplex assay were used to assay cell culture supernatant samples from human PBMCs treated with various reagents.
Over 400 Analytes to measure!
- Comprehensive Choice of Targets
- Up to 400 assays & custom formats available for human, mouse, rat, porcine, canine and primates
- Wide Range of Sample Types
- Cell culture supernatants, saliva, plasma, cell/tissue lysates, serum, BALF, pleural and peritoneal fluids & more
- Multiple Formats: 32-well and 96-well sizes available
- Key Premixed Panels
- We have designed key panels to measure Cytokines, Chemokines, Inflammation, Th1/Th2 & Th1/Th2/Th17
| Step | Protocol |
|
|
Prepare the filter plate template. Mark the standard, sample and blank wells. Standards and samples should be run in duplicates or triplicates. If the whole plate will not be used, seal the unused well with a plate seal. IMPORTANT: Place the filter plate on top of the filter plate lid during the entire assay process to prevent touching the plate bottom on any surface. |
|
2. |
Vortex working bead suspension for 15 seconds. |
|
3. |
Add 45 µL of capture bead working suspension to each well. NOTE: Save the remaining capture bead working suspension and store at 2-8°C with light protection. It can be used for setting up acquisition parameters on the flow cytometer. |
|
4. |
Remove buffer in the wells by using the “flow-through“ Filter Plate Washer connected to a vacuum source that has been adjusted according to the Filter Plate Washer Instructions. |
|
5. |
Remove buffer in the wells by using the “flow-through“ Filter Plate Washer connected to a vacuum source that has been adjusted according to the Filter Plate Washer Instructions. |
|
6. |
Add 30 µL of CCS, SPB or TL Assay Buffer to each sample well. NOTE: Cell culture supernatant samples can be run without diluting in Assay Buffer if very low levels (less than 20 pg/mL) of cytokines are expected. If it is the case, skip this step and add 45 µL of cell culture supernatant samples to each sample well in Step 7. |
|
7. |
Add 15 µL of samples to each sample well. Add 45 µL of standards to each standard well. Cover the plate with a plate seal. |
|
8. |
Incubate on the shaker (set at 700 rpm) for 60 min at room temperature. Protect from light by wrapping the filter plate in aluminum foil. |
|
9. |
Remove the plate seal. Remove solutions in the wells by using the Filter Plate Washer connected to a vacuum source. |
|
10. |
Remove solutions in the wells by using the Filter Plate Washer connected to a vacuum source. |
|
11. |
Wash the wells three times with 100µL 1x Wash Buffer using the Filter Plate Washer. |
|
12. |
Gently tap the plate bottom onto several layers of paper towels to remove residual buffer on the plate bottom after the last wash. |
|
13. |
Add 25µL of biotinylated antibody working solution to each well. Cover the plate with a plate seal. |
|
14. |
Incubate on the shaker (set at 700 rpm) for 30 min at room temperature. Protect from light by wrapping the filter plate in aluminum foil. |
|
15. |
Remove the plate seal. Remove solutions in the wells by using the Filter Plate Washer. Wash the wells three times with 100 µL 1x Wash Buffer using the Filter Plate Washer. |
|
16. |
Gently tap the plate bottom onto several layers of paper towels to remove residual buffer on the plate bottom after the last wash. |
|
17. |
Add 25µL of streptavidin-PE working solution to each well. Cover the plate with a plate seal. |
|
18. |
Incubate on the shaker (set at 700 rpm) for 20 min at room temperature. Protect from light by wrapping the filter plate in aluminum foil. |
|
19. |
Remove the plate seal. Remove solutions in the wells by using the Filter Plate Washer. Wash the wells twice with 100 µL 1x Wash Buffer. |
|
20. |
Gently tap the plate bottom onto several layers of paper towels to remove residual solution. Add 150µL to 300µL of 1x Reading Buffer to each well depending on the sample loading mechanism of a flow cytometer to re-suspend the beads. Cover the plate with a plate seal. |
|
21. |
Place the plate on the microtiter shaker and shake for 30 seconds at 700 rpm. NOTE: If the flow cytometer has no 96-well plate loader and more than 200 µL of 1x Reading Buffer is needed to re-suspend the beads, do not shake the plate. Re-suspend the beads in each well by pipetting up and down 6–8 times with a P200 pipette then transfer to a test tube for acquisition. Remove the plate seal. Read on a flow cytometer. |
| Product Code | Panel Name | Number of Analytes | Analytes |
Mouse Inflammation 5-Plex Immunoassay Panel 1 |
5 |
IL-1β, IL-6, IL-10, MCP-1, TNFα |
|
Genieplex Mouse Inflammation 5-Plex Immunoassay Panel 2 |
5 |
IL-6, IL-10, IL-12p70, MCP-1, TNFα |
|
Mouse Inflammation 6-Plex Immunoassay Panel |
6 |
IFNγ, IL-6, IL-10, IL-12p70, MCP-1, TNFα |
|
Mouse Inflammation 7-Plex Immunoassay Panel 1 |
7 |
IFNγ, IL-1β, IL-6, IP-10, KC, MCP-1, TNFα |
|
Mouse Inflammation 7-Plex Immunoassay Panel 2 |
7 |
IL-1α, IL-1β, IL-6, IL-10, IL-22, IL-23, TNFα |
|
Mouse Inflammation 8-Plex Immunoassay Panel |
8 |
IFNγ, IL-1α, IL-6, IL-10, IL-12p70, KC, MCP-1, TNFα |
|
Mouse Inflammation 11-Plex |
11 |
IFNγ, IL-1α, IL-1β, IL-6, IL-10, IL-12p70, IL-23p19, IP-10, KC, MCP-1, TNFα |
|
Mouse Inflammation 17-Plex |
17 |
IFNγ, IL-1α, IL-1β, IL-6, IL-9, IL-10, IL-12p70, IL-13, IL-15, IL-23p19, IP-10, KC, MCP-1, MIP-1α, MIP-1β, RANTES, TNFα |
| Product Code | Panel Name | Number of Analytes | Analytes |
Human T Helper Cytokine 3-Plex Immunoassay Panel 1 |
3 |
IL-9, IL-10, and TNFα |
|
Human T Helper Cytokine 3-Plex Immunoassay Panel 2 |
3 |
IL-6, IL-10, and TNFα |
|
Human T Helper Cytokine 3-Plex Immunoassay Panel 3 |
3 |
IFNγ, IL-10, and TNFα |
|
Human T Helper Cytokine 4-Plex Immunoassay Panel 1 |
4 |
IFNγ, IL-4, IL-6, TNFα |
|
Human T Helper Cytokine 4-Plex Immunoassay Panel 2 |
4 |
IFNγ, IL-4, IL-10, TNFα |
|
Human T Helper Cytokine 4-Plex Immunoassay Panel 3 |
4 |
IFNγ, IL-10, TNFα, TNFβ |
|
Human Th1/Th2 5-Plex Immunoassay Panel 1 |
5 |
IFNγ, IL-2, IL-6, IL-10, TNFα |
|
Human Th1/Th2 5-Plex Immunoassay Panel 2 |
5 |
IFNγ, IL-4, IL-6, IL-10, TNFα |
|
Human Th1/Th2 5-Plex Immunoassay Panel 3 |
5 |
IFNγ, IL-4, IL-5, IL-10, TNFα |
|
Human Th1/Th2 6-Plex Immunoassay Panel 1 |
6 |
IFNγ, IL-2, IL-4, IL-6, IL-10, TNFα |
|
Human Th1/Th2 6-Plex Immunoassay Panel 2 |
6 |
IFNγ, IL-2, IL-4, IL-5, IL-10, TNFα |
|
Human Th1/Th2 7-Plex Immunoassay Panel |
7 |
IFNγ, IL-2, IL-4, IL-5, IL-6, IL-10, TNFα |
|
Human Th1/Th2 10-plex |
10 |
IFNγ, IL-1beta, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, TNFα |
|
Human Th1/Th2 11-plex |
11 |
GM-CSF, IFNγ, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-9, IL-10, IL-12p70, TNFα |
|
Human T Helper Cytokine 14-Plex Immunoassay Panel |
14 |
GM-CSF, IFNγ, IL-2, IL-4, IL-5, IL-6, IL-9, IL-10, IL-12p70, IL-13, IL-17A, IL-21, IL-22 and TNFα |
|
Human Th1/Th2/Th17 7-plex Immunoassay Panel |
7 |
IFNγ, IL-2, IL-4, IL-6, IL-10, IL-17A, TNFα |
|
Human Th1/Th2/Th17 8-plex Immunoassay Panel 1 |
8 |
IFNγ, IL-2, IL-4, IL-6, IL-8, IL-10, IL-17A, TNFα |
|
Human Th1/Th2/Th17 11-plex |
11 |
IFNγ, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-10, IL-12p70, IL-17A, TNFα, TNFβ |
|
Human Th1/Th2/Th17 12-plex |
12 |
IFNγ, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-17A, TNFα, TNFβ |
|
Human Th1/Th2/Th17 14-plex |
14 |
IFNγ, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-17A, IL-17F, IL-22, TNFα, TNFβ |
|
Human Th1/Th2/Th17 18-plex |
18 |
GM-CSF, IFNγ, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-8, IL-9, IL-10, IL-12p70, IL-17A, IL-17F, IL-22, IL-33, TNFα, TNFβ, TSLP |
|
Human Th1/Th2/Th17/Treg 10-Plex Immunoassay Panel |
10 |
G-CSF, GM-CSF, IFNγ, IL-2, IL-4, IL-6, IL-10, IL-17A, TNFα and TNFβ |
|
Human Inflammation 6-Plex Immunoassay Panel |
6 |
IL-1β, IL-6, IL-8, IL-10, IL-12p70, TNFα |
|
Human Inflammation 7-Plex Immunoassay Panel |
7 |
IL-1β, IL-1RA, IL-6, IL-8, IL-10, IL-12p70, TNFα |
|
Human Inflammation 11-Plex Immunoassay Panel |
11 |
IFNγ, IL-1α, IL-1β, IL-6, IL-8, IL-10, IL-12p70, IL-27, IP-10, MCP-1, TNFα |
|
Human Inflammation 16-Plex Immunoassay Panel |
16 |
IFNγ, IL-1α, IL-1β, IL-6, IL-8, IL-10, IL-12p70, IL-13, IL-17A, IL-27, IL-31, IL-33, IP-10, MCP-1, MIP-1α, and TNFα |
|
Human Inflammation 18-Plex Immunoassay Panel |
18 |
IFNγ, IL-1α, IL-1β, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, IL-17A, IL-27, IL-31, IL-33, IP-10, MCP-1, MIP-1α, MIP-1β, and TNFα |
|
Human Chemokine 6-Plex Immunoassay Panel |
6 |
Eotaxin, IL-8, RANTES, MCP-1, MCP-3, IP-10 |
|
Human Inflammatory Chemokine 7-Plex Immunoassay Panel 1 |
7 |
BCA-1, MCP-1, MIP-1α, MIP-3β, PARC, PF4, RANTES |
|
Human Inflammatory Chemokine 7-Plex Immunoassay Panel 2 |
7 |
IP-10, I-TAC, MIP-1α, MIP-1β, 6Ckine, MDC, MIP-3β |
|
Human CD8+ T Cell Related Cytokines, 14-plex Immunoassay Panel |
14 |
GM-CSF, Granzyme B, IFNγ, IL-2, IL-4. IL-5, IL6, IL-10, IL-13, IL-15, MIP-1α, MIP-1β, sFasL, TNFα |
|
Human CD8+ T Cell Related Cytokines, 16-plex Immunoassay Panel |
16 |
GM-CSF, Granzyme B, IFNγ, IL-2, IL-4. IL-5, IL-6, IL-10, IL-13, IL-15, MIP-1α, MIP-1β, RANTES, sCD137, sFasL, TNFα |
| Product Code | Panel Name | Number of Analytes | Analytes |
Mouse T Helper Cytokine 3-Plex Immunoassay Panel 1 |
3 |
IL-9, IL-10, and TNFα |
|
Mouse T Helper Cytokine 3-Plex Immunoassay Panel 2 |
3 |
IFN-Gamma, IL-10, and TNF-Alpha |
|
Mouse T Helper Cytokine 3-Plex Immunoassay Panel 3 |
3 |
IL-9, IL-10, and TNFα |
|
Mouse T Helper Cytokine 4-Plex Immunoassay Panel 1 |
4 |
IFNγ, IL-4, IL-6, TNFα |
|
Mouse T Helper Cytokine 4-Plex Immunoassay Panel 2 |
4 |
IFNγ, IL-4, IL-10, TNFα |
|
Mouse T Helper Cytokine 4-Plex Immunoassay Panel 3 |
4 |
IL-6, IL-10, IL-22, TNFα |
|
Mouse Th1/Th2 5-plex Immunoassay Panel |
5 |
IFNγ, IL-2, IL-4, IL-5, TNFα |
|
Mouse Th1/Th2 6-plex Immunoassay Panel |
6 |
IFNγ, IL-2, IL-4, IL-6, IL-10, TNFα |
|
Mouse Th1/Th2 6-plex Immunoassay Panel |
6 |
IFNγ, IL-2, IL-4, IL-5, IL-10, TNFα |
|
Mouse Th1/Th2 8-Plex Immunoassay Panel |
8 |
IFNγ, IL-1α, IL-2, IL-4, IL-5, IL-6, IL-10, and TNFα |
|
Mouse Th1/Th2/Th17 7-plex Kit |
7 |
IFNγ, IL-2, IL-4, IL-6, IL-10, IL-17A, TNFα |
|
Mouse Th1/Th2/Th17 10-plex Kit |
10 |
GM-CSF, IFNγ, IL-1α, IL-2, IL-4, IL-5, IL-6, IL-10, IL-17A, TNFα |
|
Mouse Th1/Th2/Th17 14-plex |
14 |
IFNγ, IL-1α, IL-1β, IL-2 , IL-4, IL-5, IL-6, IL-10, IL-12p70, IL-13, IL-17A, IL-21, IL-22, TNFα |
|
Mouse Th1/Th2/Th17 16-plex |
16 |
GM-CSF, IFNγ, IL-1α, IL-1β, IL-2 , IL-4, IL-5, IL-6, IL-10, IL-12p70, IL-13, IL-17A, IL-21, IL-23p19, KC, TNFα |
|
Mouse Th1/Th2/Th17 18-plex |
18 |
GM-CSF, IFNγ, IL-1α, IL-1β, IL-2 , IL-4, IL-5, IL-6, IL-10, IL-12p70, IL-13, IL-17A, IL-21, IL-22, IL-23p19, KC, TNFα, TSLP |
|
Mouse Inflammation 5-Plex Immunoassay Panel 1 |
5 |
IL-1β, IL-6, IL-10, MCP-1, TNFα |
|
Genieplex Mouse Inflammation 5-Plex Immunoassay Panel 2 |
5 |
IL-6, IL-10, IL-12p70, MCP-1, TNFα |
|
Mouse Inflammation 6-Plex Immunoassay Panel |
6 |
IFNγ, IL-6, IL-10, IL-12p70, MCP-1, TNFα |
|
Mouse Inflammation 7-Plex Immunoassay Panel 1 |
7 |
IFNγ, IL-1β, IL-6, IP-10, KC, MCP-1, TNFα |
|
Mouse Inflammation 7-Plex Immunoassay Panel 2 |
7 |
IL-1α, IL-1β, IL-6, IL-10, IL-22, IL-23, TNFα |
|
Mouse Inflammation 8-Plex Immunoassay Panel |
8 |
IFNγ, IL-1α, IL-6, IL-10, IL-12p70, KC, MCP-1, TNFα |
|
Mouse Inflammation 11-Plex |
11 |
IFNγ, IL-1α, IL-1β, IL-6, IL-10, IL-12p70, IL-23p19, IP-10, KC, MCP-1, TNFα |
|
Mouse Inflammation 17-Plex |
17 |
IFNγ, IL-1α, IL-1β, IL-6, IL-9, IL-10, IL-12p70, IL-13, IL-15, IL-23p19, IP-10, KC, MCP-1, MIP-1α, MIP-1β, RANTES, TNFα |
|
Mouse Chemokine 7-Plex Immunoassay Panel |
7 |
Eotaxin, KC, MCP-1, MCP-3, MIG, RANTES, IP-10 |