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Rat androgen receptor(AR) ELISA Kit

96 Tests
Catalogue Number:SL0824Ra

Store all reagents at 2-8℃

Validity Perid: six months

For samples:

In serum, plasma, culture media or any biological fluid.

FOR RESEARCH USE ONLY！

NOT FOR THERAPEUTIC OR DIAGNOSTIC APPLICATIONS！

PLEASE READTHROUGH ENTIRE PROCEDURE BEFORE BEGINNING

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Rat androgen receptor(AR) ELISA Kit
FOR RESEARCH USE ONLY

Drug Names
Generic Name：Rat androgen receptor(AR) ELISA Kit

Purpose
Our Rat androgen receptor(AR) ELISA Kit is to assay AR levels in Rat serum, plasma,
culture media or any biological fluid.

Principle
This ELISA kit uses Sandwich-ELISA as the method. The Microelisa stripplate provided
in this kit has been pre-coated with an antibody specific to AR. Standards or samples are
added to the appropriate Microelisa stripplate wells and combined to the specific antibody.
Then a Horseradish Peroxidase (HRP)- conjugated antibody specific for AR is added to each
Microelisa stripplate well and incubated. Free components are washed away. The TMB
substrate solution is added to each well. Only those wells that contain AR and HRP
conjugated AR antibody will appear blue in color and then turn yellow after the addition of
the stop solution. The optical density (OD) is measured spectrophotometrically at a
wavelength of 450 nm. The OD value is proportional to the concentration of AR. You can
calculate the concentration of AR in the samples by comparing the OD of the samples to the
standard curve.

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Materials provided with the kit
Materials provided with the kit 96 determinations Storage
1 User manual 1 R.T.
2 Closure plate membrane 2 R.T.
3 Sealed bags 1 R.T.
4 Microelisa stripplate 1 2-8℃
5 Standard：27 ng/ml 0.5ml×1 bottle 2-8℃
6 Standard diluent 1.5ml×1 bottle 2-8℃
7 HRP-Conjugate reagent 6ml×1 bottle 2-8℃
8 Sample diluent 6ml×1 bottle 2-8℃
9 Chromogen Solution A 6ml×1 bottle 2-8℃
10 Chromogen Solution B 6ml×1 bottle 2-8℃
11 Stop Solution 6ml×1 bottle 2-8℃
12 wash solution 20ml (30X)×1bottle 2-8℃

Sample preparation
1. Serum preparation
After collection of the whole blood, allow the blood to clot by leaving it undisturbed at
room temperature. This usually takes 10-20 minutes. Remove the clot by centrifuging at
2,000-3,000 rpm for 20 minutes. If precipitates appear during reservation, the sample should
be centrifugated again.
2. Plasma preparation
Collect the whole blood into tubes with anticoagulant (EDTA or citrate). After incubated at
room temperature for 10-20 minutes, tubes are centrifugated for 20 min at 2,000-3,000 rpm.
Collect the supernatant carefully as plasma samples. If precipitates appear during reservation,
the sample should be centrifugated again.
3. Urine samples
Collect urine into aseptic tubes. Collect the supernatant carefully after centrifuging for 20
min at 2,000-3,000 rpm. If precipitates appear during reservation, the sample should be
centrifugated again. The preparation procedure of cerebrospinal fluid and pleuroperitoneal
fluid is the same as that of urine sample.
4. Cell samples
If you want to detect the secretions of cells, collect culture supernatant into aseptic tubes.

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Collect the supernatant carefully after centrifuging for 20 min at 2,000-3,000 rpm. If you want
to detect intracellular components, dilute the cells to 1X100/ml with PBS (pH 7.2-7.4). The
cells were destroyed to release intracellular components by repeated freezing and thawing.
Collect the supernatant carefully after centrifuging for 20 min at 2,000-3,000 rpm. If
precipitates appear during reservation, the sample should be centrifugated again.
5. Tissue samples
Tissue samples are cut, weighed, frozen in liquid nitrogen and stored at -80℃ for future
use. The tissue samples were homogenized after adding PBS (pH 7.4). Samples should be
operated at 4℃. Collect the supernatant carefully after centrifuging for 20 min at 2,000-3,000
rpm. Aliquot the supernatant for ELISA assay and future use.
Notes:
1. Sample extraction and ELISA assay should be performed as soon as possible after sample
collection. The samples should be extracted according to the relevant literature. If ELISA
assay can not be performed immediately, samples can be stored at -20 ℃ .Repeated
freeze-thaw cycles should be avoided.
2. Our kits can not be used for samples with NaN3 which can inhibit the activity of HRP.

Procedure
1. Dilution of Standards
Ten wells are set for standards in a Microelisa stripplate. In Well 1 and Well 2, 100μl
Standard solution and 50μl Standard Dilution buffer are added and mixed well. In Well 3
and Well 4, 100μl solution from Well 1 and Well 2 are added respectively. Then 50μl
Standard Dilution buffer are added and mixed well. 50μl solution is discarded from Well 3
and Well 4. In Well 5 and Well 6, 50μl solution from Well 3 and Well 4 are added
respectively. Then 50μl Standard Dilution buffer are added and mixed well. In Well 7 and
Well 8, 50μl solution from Well 5 and Well 6 are added respectively. Then 50μl Standard
Dilution buffer are added and mixed well. In Well 9 and Well 10, 50μl solution from Well
7 and Well 8 are added respectively. Then 50μl Standard Dilution buffer are added and
mixed well. 50μl solution is discarded from Well 9 and Well 10. After dilution, the total

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volume in all the wells are 50μl and the concentrations are 18 ng/ml, 12 ng/ml, 6 ng/ml, 3
ng/ml and 1.5 ng/ml, respectively.

27 ng/ml 18 ng/ml 12 ng/ml 6 ng/ml 3 ng/ml 1.5 ng/ml

2. In the Microelisa stripplate, leave a well empty as blank control. In sample wells, 40μl
Sample dilution buffer and 10μl sample are added (dilution factor is 5). Samples should be
loaded onto the bottom without touching the well wall. Mix well with gentle shaking.
3. Incubation: incubate 30 min at 37℃ after sealed with Closure plate membrane.
4. Dilution: dilute the concentrated washing buffer with distilled water (30 times for 96T and
20 times for 48T).
5. Washing: carefully peel off Closure plate membrane, aspirate and refill with the wash
solution. Discard the wash solution after resting for 30 seconds. Repeat the washing
procedure for 5 times.
6. Add 50 μl HRP-Conjugate reagent to each well except the blank control well.
7. Incubation as described in Step 3.
8. Washing as described in Step 5.
9. Coloring: Add 50 μl Chromogen Solution A and 50 μl Chromogen Solution B to each
well, mix with gently shaking and incubate at 37 ℃ for 15 minutes. Please avoid light
during coloring.
10. Termination: add 50 μl stop solution to each well to terminate the reaction. The color in
the well should change from blue to yellow.
11. Read absorbance O.D. at 450nm using a Microtiter Plate Reader. The OD value of the
blank control well is set as zero. Assay should be carried out within 15 minutes after

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adding stop solution.

Notes:
1．Store the kit at 4 ° C upon receipt.The kit should be equilibrat4ed to room temperature
before the assay. Remove any unneeded strips from Rat AR Antibody-Coated plate, reseal
them in zip-lock foil and keep at 4°C.
2．Precipitates may appear in concentrated washing buffer. Please heat the buffer to dissolve
all the precipitates, which will not affect the results.
3．Accurate pipette should be used to avoid experimental error. Samples should be added to
the Microplate in less than 5 minutes. If a large number of samples are included, multiple
channel pipette is recommended.
4．Standard curve should be included in every assay. Replicate wells are recommended. If
the OD value of the sample is greater than the first well of standards, please dilute the
sample (n times) before test. When calculating the original AR concentration, please
multiply the total dilution factor (XnX5).
5．In order to avoid cross-contamination, Microplate sealers are for one-time use only.
6．Please keep Substrate away from light.
7．All the operation should be accordance with the manufacturer's instructions strictly. The
results determined by the Microtiter Plate Reader.
8．All the samples, washing buffer and wastes should be treated as infectious agents.
9．Reagents from different lots should not be mixed.

Calculation of Results
Known concentrations of Rat AR Standard and its
corresponding reading OD is plotted on the log scale
(x-axis) and the log scale (y-axis) respectively. The
concentration of Rat AR in sample is determined by
plotting the sample’s O.D. on the Y-axis. The original
concentration is calculated by multiplying the dilution
factor. This diagram is for reference only

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Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level
Rat AR were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, middle and high level
Rat AR were tested on 3 different plates, 8 replicates in each plate.
CV(%) = SD/meanX100
Intra-Assay: CV<10%
Inter-Assay: CV<12%

Assay range
0.3ng/ml - 20 ng/ml

Sensitivity：
0.05 ng/ml

Storage and validity

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