Customized Human METRNL ELISA Assay Kit Meteorin-like Protein ELISA
Test Kit For Research
Cat.No E3941Hu
Size: 96 wells
Storage: Store the reagents at 2-8°C. For over 6-month storage refer to
the expiration date keep it at -20°C. Avoid repeated thaw cycles.
If individual reagents are opened it is recommended that the kit be
used within 1 month.
*This product is for research use only, not for use in diagnosis
procedures. It’s highly recommend to read this instruction entirely
before use.
Intended Use
This ELISA kit applies to the invitro quantitative determination of
human METRNL concentrations in serum, plasma, tissue and other
biological fluids.
Precision
Intra-Assay Precision (Precision within an assay) Three samples of
known concentration were tested on one plate to assess intra-assay
precision.
Inter-Assay Precision (Precision between assays) Three samples of
known concentration were tested in separate assays to assess
inter-assay precision.
CV(%) = SD/mean x 100
Intra-Assay: CV<8%
Inter-Assay: CV<10%
Reagent Provided
Components | Quantity |
Standard Solution (16ng/ml) | 0.5ml x1 |
Pre-coated ELISA Plate | 12 * 8 well strips x1 |
Standard Diluent | 3ml x1 |
Streptavidin-HRP | 6ml x1 |
Stop Solution | 6ml x1 |
Substrate Solution A | 6ml x1 |
Substrate Solution B | 6ml x1 |
Wash Buffer Concentrate (25x) | 20ml x1 |
Biotinylated human METRNL Antibody | 1ml x1 |
User Instruction | 1 |
Plate Sealer | 2 pics |
Zipper bag | 1 pic |
Specimen Collection
Samples should be clear and transparent and be centrifuged to
remove suspended solids.
Serum: Allow samples to clot for 2 hours at room temperature or overnight
at 4°C before centrifugation for 15 minutes at 1000×g. Collect the
supernatant and carry out the assay immediately. Blood collection
tubes should be disposable, non-pyrogenic, and non-endotoxin.
Plasma: Collect plasma using EDTA or heparin as an anticoagulant.
Centrifuge samples for 15 minutes at 1000×g at 2 - 8°Cwithin 30
minutes of collection. Collect the supernatant and carry out the
assay immediately. Hemolysissamples are not suitable for ELISA
assay!
Cell culture supernate: Centrifuge supernate for 20 minutes to remove insoluble impurity
and cell debris at 1000×g at 2 - 8°C. Collect the clear supernate
and carry out the assay immediately.
Tissue homogenates:You’d better get detailed references from other literatures before
assay aiming at different tissue types. For general information,
hemolysis blood may affect the result, so you should mince the
tissues to small pieces and rinse them in ice-cold PBS (0.01M,
pH=7.4) to remove excess blood thoroughly. Tissue pieces should be
weighed and then homogenized in PBS (the volume depends on the
weight of the tissue) with a glass homogenizer on ice. To further
break the cells, you can sonicate the suspension with an ultrasonic
cell disrupter or subject it to freeze-thaw cycles. The homogenates
are then centrifugated for 5minutes at 5000×g to get the supernate.
Other biological fluids: Centrifuge samples for 20 minutes at 1000×g at 2 - 8°C. Collect
the supernatant and carry out the assay immediately.
Note
- Sample concentrations should be predicted before being used in the
assay. If the sample concentration is not within the range of the
standard curve, users must contact us to determine the optimal sample for their particular experiments.
- Samples to be used within 5 days should be stored at 2-8°C. Samples
should be aliquoted or must be stored at -20°C within 1 month or
-80°C within 6 months. Avoid repeated freeze thaw cycles.
- Samples should be brought to room temperature before starting the
assay.
- Centrifuge to collect sample before use.
- Samples containing NaN3 can’t be tested as it inhibits the activity
of Horse Radish Peroxidase (HRP).
- Collect the supernatants carefully. When sediments occurred during
storage, centrifugation should be performed again.
- Hemolysis can greatly impact the validity of test results. Take
care to minimize hemolysis.
*Sample can't be diluted with this kit. Owing to the the material we use to prepare the kit, the sample
matrix interference may falsely depress the specificity and
accuracy of the assay.
Assay Procedure
1. Prepare all reagents, standard solutions and samples as
instructed. Bring all reagents to room temperature before use. The
assay is performed at room temperature.
2. Determine the number of strips required for the assay. Insert
the strips in the frames for use. The unused strips should be
stored at 2-8°C.
3. Add 50μl standard to standard well. Note: Don’t add antibody to standard well because the standard solution
contains biotinylated antibody.
4. Add 40μl sample to sample wells and then add 10μl anti-METRNL
antibody to sample wells, then add 50μl streptavidin-HRP to sample
wells and standard wells (Not blank control well). Mix well. Cover
the plate with a sealer. Incubate 60 minutes at 37°C.
5. Remove the sealer and wash the plate 5 times with wash buffer.
Soak wells with at least 0.35 ml wash buffer for 30 seconds to 1
minute for each wash. For automated washing, aspirate all wells and
wash 5 times with wash buffer, overfilling wells with wash buffer.
Blot the plate onto paper towels or other absorbent material.
6. Add 50μl substrate solution A to each well and then add 50μl
substrate solution B to each well. Incubate plate covered with a
new sealer for 10 minutes at 37°C in the dark.
7. Add 50μl Stop Solution to each well, the blue color will change
into yellow immediately.
8. Determine the optical density (OD value) of each well
immediately using a microplate reader set to 450 nm within 10
minuets after adding the stop solution.
Calculation of Result
Construct a standard curve by plotting the average OD for each
standard on the vertical (Y) axis against the concentration on the
horizontal (X) axis and draw a best fit curve through the points on
the graph. These calculations can be best performed with
computer-based curve-fitting software and the best fit line can be
determined by regression analysis.