Rat RF-IgG (Rheumatoid Factor IgG) ELISA Kit (RTFI01409)

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SKU:
RTFI01409
Product Type:
ELISA Kit
Size:
96 Assays
Sensitivity:
1.875U/ml
Range:
3.125-200IU/ml
ELISA Type:
Indirect ELISA - Biotin
Synonyms:
Rheumatoid Factor IgG, RF-IgG
Reactivity:
Rat
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Description

system_update_altTechnical Manual

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商品名:Rat RF-IgG (Rheumatoid Factor IgG) ELISA Kit
製品コード:RTFI01409
サイズ:96 Assays
目標:Rat RF-IgG (Rheumatoid Factor IgG)
エイリアス:Rheumatoid Factor IgG ELISA Kit, RF-IgG ELISA Kit
反応性:Rat
検出方法:Indirect ELISA - Biotin
感度:< 1.875U/ml
範囲:3.125-200IU/ml
保管所:4°C for 6 months
ノート:For Research Use Only
回復:Matrices listed below were spiked with certain level of Rat RF-IgG (Rheumatoid Factor IgG) and the recovery rates were calculated by comparing the measured value to the expected amount of Rat RF-IgG (Rheumatoid Factor IgG) in samples.
MatrixRecovery range(%)Average(%)
serum(n=5)85-10596
EDTA plasma(n=5)87-10295
heparin plasma(n=5)88-9992
直線性:The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Rat RF-IgG (Rheumatoid Factor IgG) and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.
Sample1:21:41:8
serum(n=5)85-105%85-101%89-105%
EDTA plasma(n=5)82-101%85-92%85-94%
UFH plasma(n=5)84-99%81-97%83-100%
Intra-Assay:CV <8%
Inter-Assay:CV <10%
ステッププロトコル
1. Set standard, test sample (diluted at least 50% with Sample Dilution Buffer) and control (blank) wells on the pre-coated plate and record their positions. It is recommended to measure each standard and sample in duplicate. Wash plate 2 times before adding standard, sample and control (blank) wells!
2. Aliquot 100µL of standard solutions into the standard wells.
3. Add 100µL of properly diluted Sample into the sample wells.
4. Seal the plate with a cover and incubate at 37°C for 90 mins.
5. Wash: Repeat the aspiration/wash process for 2 times.
6. Add 100µL of Biotin-labelled antibody working solution into the bottom of each well (standard, test sample & zero wells) without touching the side walls.
7. Seal the plate with a cover and incubate at 37°C for 60 mins.
8. Wash: Remove the cover, and wash plate 3 times with Wash buffer.
9. HRP-Streptavidin Conjugate (SABC): Add 100µL of SABC Working Solution into each well, cover the plate and incubate at 37°C for 30 minutes.
10. Wash: Remove the cover and wash plate 5 times with Wash Buffer.
11. TMB Substrate: Add 90 µL of TMB substrate into each well, cover the plate and incubate at 37°C in dark for 10-20 mins. (Note: This incubation time is for reference only, the optimal time should be determined by the end-user.) As soon as a blue colour develops in the first 3-4 wells (with most concentrated standards) and the other wells show no obvious colour, terminate the reaction.
12. Add 50µL of Stop solution into each well and mix thoroughly. The colour changes into yellow immediately.
13. Read the O.D. absorbance at 450 nm in a microplate reader immediately after adding the stop solution.

ELISAアッセイを実施する場合、可能な限り最良の結果を達成するためにサンプルを準備することが重要です。以下に、さまざまなサンプルタイプのサンプルを準備するための手順のリストを示します。

サンプルタイプ プロトコル
血清

If using serum separator tubes, allow samples to clot for 30 minutes at room temperature. Centrifuge for 10 minutes at 1,000x g. Collect the serum fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.

If serum separator tubes are not being used, allow samples to clotovernight at 2-8°C. Centrifuge for 10 minutes at 1,000x g. Removeserum and assay promptly or aliquot and store the samples at-80°C. Avoid multiple freeze-thaw cycles.

プラズマ Collect plasma using EDTA or heparin as an anti-coagulant. Centrifuge samples at 4°C for 15 mins at 1000 × g within 30 mins of collection. Collect the plasma fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.Note: Over haemolysed samples are not suitable for use with this kit.
尿および脳脊髄液 Collect the urine (mid-stream) in a sterile container, centrifuge for 20 mins at 2000-3000 rpm. Remove supernatant and assay immediately. If any precipitation is detected, repeat the centrifugation step. A similar protocol can be used for cerebrospinal fluid.
細胞培養上清 Collect the cell culture media by pipette, followed by centrifugation at 4°C for 20 mins at 1500 rpm. Collect the clear supernatant and assay immediately.
細胞溶解物 Solubilize cells in lysis buffer and allow to sit on ice for 30 minutes. Centrifuge tubes at 14,000 x g for 5 minutes to remove insoluble material. Aliquot the supernatant into a new tube and discard the remaining whole cell extract. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20°C.
組織ホモジネート The preparation of tissue homogenates will vary depending upon tissue type. Rinse tissue with 1X PBS to remove excess blood & homogenizein 20ml of 1X PBS (including protease inhibitors) and store overnight at ≤ -20°C. Two freeze-thaw cycles are required to break the cell membranes. To further disrupt the cell membranes you can sonicate the samples. Centrifuge homogenates for 5 mins at 5000xg. Remove the supernatant and assay immediately or aliquot and store at -20°C or-80°C.
組織溶解物 Rinse tissue with PBS, cut into 1-2 mm pieces, and homogenize with a tissue homogenizer in PBS. Add an equal volume of RIPA buffer containing protease inhibitors and lyse tissues at room temperature for 30 minutes with gentle agitation. Centrifuge to remove debris. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C.
母乳 Collect milk samples and centrifuge at 10,000 x g for 60 min at 4°C. Aliquot the supernatant and assay. For long term use, store samples at -80°C. Minimize freeze/thaw cycles.
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