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Human Apoptosis Inducing Factor (AIF) ELISA Kit
two product lines: Traditional ELISA Kit and Ready-to-Use ELISA Kit.
| Traditional ELISA Kit | Ready-to-Use ELISA KIT | |
| Product name: | Human Apoptosis Inducing Factor (AIF) ELISA Kit | |
| Method: | Sandwich | |
| Synonyms: | PDCD8; Programmed Cell Death protein 8; Apoptosis-inducing factor 1, mitochondrial |
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| Detection range: | 0.156-10ng/mL | |
| Target Protein: | AIF | |
| Size: | 96T/48T | |
| Quality guarantee period: | for 12 months, 16 months | |
| Catalog number: | DL-AIF-Hu (traditional) | (ready-to-use) |
| Assay length | 1-4.5Hours | 1-3.5Hours |
| Advantages: |
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| Instruction Manual |
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Other names：PDCD8; Programmed Cell Death protein 8; Apoptosis-inducing factor 1, mitochondrial
Function: Probable oxidoreductase that has a dual role in controlling cellular life and death; during apoptosis, it is translocated from the mitochondria to the nucleus to function as a proapoptotic factor in a caspase-independent pathway, while in normal mitochondria, it functions as an antiapoptotic factor via its oxidoreductase activity. The soluble form (AIFsol) found in the nucleus induces 'parthanatos' i.e., caspase-independent fragmentation of chromosomal DNA. Interacts with EIF3G,and thereby inhibits the EIF3 machinery and protein synthesis, and activates casapse-7 to amplify apoptosis. Plays a critical role in caspase-independent, pyknotic cell death in hydrogen peroxide-exposed cells. Binds to DNA in a sequence-independent manner.
Sequence：
10 20 30 40 50 MFRCGGLAAG ALKQKLVPLV RTVCVRSPRQ RNRLPGNLFQ RWHVPLELQM 60 70 80 90 100 TRQMASSGAS GGKIDNSVLV LIVGLSTVGA GAYAYKTMKE DEKRYNERIS 110 120 130 140 150 GLGLTPEQKQ KKAALSASEG EEVPQDKAPS HVPFLLIGGG TAAFAAARSI 160 170 180 190 200 RARDPGARVL IVSEDPELPY MRPPLSKELW FSDDPNVTKT LRFKQWNGKE 210 220 230 240 250 RSIYFQPPSF YVSAQDLPHI ENGGVAVLTG KKVVQLDVRD NMVKLNDGSQ 260 270 280 290 300 ITYEKCLIAT GGTPRSLSAI DRAGAEVKSR TTLFRKIGDF RSLEKISREV 310 320 330 340 350 KSITIIGGGF LGSELACALG RKARALGTEV IQLFPEKGNM GKILPEYLSN 360 370 380 390 400 WTMEKVRREG VKVMPNAIVQ SVGVSSGKLL IKLKDGRKVE TDHIVAAVGL 410 420 430 440 450 EPNVELAKTG GLEIDSDFGG FRVNAELQAR SNIWVAGDAA CFYDIKLGRR 460 470 480 490 500 RVEHHDHAVV SGRLAGENMT GAAKPYWHQS MFWSDLGPDV GYEAIGLVDS 510 520 530 540 550 SLPTVGVFAK ATAQDNPKSA TEQSGTGIRS ESETESEASE ITIPPSTPAV 560 570 580 590 600 PQAPVQGEDY GKGVIFYLRD KVVVGIVLWN IFNRMPIARK IIKDGEQHED 610 LNEVAKLFNI HED
INTENDED USE
The kit is a sandwich enzyme immunoassay for the in vitro quantitative measurement of AIF in human serum, plasma, tissue homogenates or other biological fluids.
DETECTION RANGE
0.156-10ng/mL. The standard curve concentrations used for the ELISA’s were 10ng/mL, 5ng/mL, 2.5ng/mL, 1.25ng/mL, 0.625ng/mL, 0.312ng/mL, 0.156ng/mL.
SENSITIVITY
The minimum detectable dose of AIF is typically less than 0.064ng/mL.
The sensitivity of this assay, or Lower Limit of Detection (LLD) was defined as the lowest protein concentration that could be differentiated from zero. It was determined by adding two standard deviations to the mean optical density value of twenty zero standard replicates and calculating the corresponding concentration.
SPECIFICITY
This assay has high sensitivity and excellent specificity for detection of AIF.
No significant cross-reactivity or interference between AIF and analogues was observed.
You can reference link of the kit as following
https://dldevelop.com/Research-reagent/dl-aif-hu.html
https://dldevelop.com/Research-reagent/dl-aif-ra.html
Introduction
| Item | Standard | Test | |
| Description |
The kit is a sandwich enzyme immunoassay for the in vitro quantitative measurement of AIF in human serum, plasma, tissue homogenates and other biological fluids. |
Conform | |
| Identification | Colorimetric | Positive | |
| Composition | Traditional ELISA Kit | Ready-to-Use ELISA KIT | Conform |
| Pre-coated, ready to use 96-well strip plate 1 | Pre-coated, ready to use 96-well strip plate 1 | ||
| Plate sealer for 96 wells 2 | Plate sealer for 96 wells 2 | ||
| Standard 2 | Standard 2 | ||
| Diluents buffer 1×45mL | Standard Diluent 1×20mL | ||
| Detection Reagent A 1×120μL | Detection Solution A 1×12mL | ||
| Detection Reagent B 1×120μL | Detection Solution B 1×12mL | ||
| TMB Substrate 1×9mL | TMB Substrate 1×9mL | ||
| Stop Solution 1×6mL | Stop Solution 1×6mL | ||
| Wash Buffer (30 × concentrate) 1×20mL | Wash Buffer (30 × concentrate) 1×20mL | ||
| Instruction manual 1 | Instruction manual 1 | ||
Test principle
The microtiter plate provided in this kit has been pre-coated with an antibody specific to the index. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific to the index. Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After TMB substrate solution is added, only those wells that contain the index, biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450nm ± 10nm. The concentration of the index in the samples is then determined by comparing the O.D. of the samples to the standard curve.
Recovery
Matrices listed below were spiked with certain level of recombinant AIF and the recovery rates were calculated by comparing the measured value to the expected amount of the index in samples.
| Matrix | Recovery range (%) | Average(%) |
| serum(n=5) | 81-93 | 86 |
| EDTA plasma(n=5) | 80-97 | 88 |
| heparin plasma(n=5) | 90-101 | 95 |
Linearity
The linearity of the kit was assayed by testing samples spiked with appropriate concentration of the index and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.
| Sample | 1：2 | 1：4 | 1：8 | 1：16 |
| serum(n=5) | 82-96% | 83-98% | 81-99% | 93-101% |
| EDTA plasma(n=5) | 88-101% | 86-95% | 90-102% | 80-93% |
| heparin plasma(n=5) | 80-91% | 82-90% | 95-104% | 79-95% |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level the index were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, middle and high level the index were tested on 3 different plates, 8 replicates in each plate.
CV(%) = SD/meanX100
Intra-Assay: CV<10%
Inter-Assay: CV<12%
Stability
The stability of ELISA kit is determined by the loss rate of activity. The loss rate of this kit is less than 5% within the expiration date under appropriate storage conditions.
Note:
To minimize unnecessary influences on the performance, operation procedures and lab conditions, especially room temperature, air humidity and incubator temperatures should be strictly regulated. It is also strongly suggested that the whole assay is performed by the same experimenter from the beginning to the end.
Assay procedure summary
1. Prepare all reagents, samples and standards;
2. Add 100µL standard or sample to each well. Incubate 2 hours at 37℃;
3. Aspirate and add 100µL prepared Detection Reagent A. Incubate 1 hour at 37℃;
4. Aspirate and wash 3 times;
5. Add 100µL prepared Detection Reagent B. Incubate 1 hour at 37℃;
6. Aspirate and wash 5 times;
7. Add 90µL Substrate Solution. Incubate 15-25 minutes at 37℃;
8. Add 50µL Stop Solution. Read at 450nm immediately.
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