FTH1 ELISA Kit

Catalog No. ABIN579138

Product Details FTH1 ELISA Kit

Target: FTH1 (Ferritin, Heavy Polypeptide 1 (FTH1))

Reactivity: Rat

Detection Method: Colorimetric

Method Type: Sandwich ELISA

Detection Range: 6.25-400 ng/mL

Minimum Detection Limit: 6.25 ng/mL

Application: ELISA

Purpose: This immunoassay kit allows for the specific measurement of Rat Ferritin concentrations in serum and plasma.

Sample Type: Serum, Plasma

Analytical Method: Quantitative

Specificity: This assay recognizes recombinant and natural Rat Ferritin.

Cross-Reactivity (Details): No significant cross-reactivity or interference was observed.

Characteristics: Rattus norvegicus,Rat,Ferritin heavy chain,Ferritin H subunit,Fth1,Fth,1.16.3.1

Components: Reagent (Quantity ): Assay plate (1), Standard 2 Sample Diluent (1 × 20ml), Assay Diluent A (1x10ml), Assay Diluent B (1x10ml), Detection Reagent A (1 × 120μl), 2 Detection Reagent B 1 x 120μl Wash Buffer (25 x concentrate) (1 × 30ml), Substrate (1 x 10ml) Stop Solution (1 x 10ml)

Application Details

Sample Volume: 100 μL

Plate: Pre-coated

Protocol: This assay employs the quantitative sandwich enzyme immunoassay technique. A monoclonal antibody specific for Ferritin has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any Ferritin present is bound by the immobilized antibody. An enzyme-linked monoclonal antibody specific for Ferritin is added to the wells. Following a wash to remove any unbound antibody-enzyme reagent, a substrate solution is added to the wells and color develops in proportion to the amount of Ferritin bound in the initial step. The color development is stopped and the intensity of the color is measured.

Reagent Preparation:

Bring all reagents to room temperature before use. Wash Buffer - If crystals have formed in the concentrate, warm to room temperature and mix gently until the crystals have completely dissolved. Dilute 30 mL of Wash Buffer Concentrate into deionized or distilled water to prepare 750 mL of Wash Buffer. Standard - Reconstitute the Standard with 1.0 ml of Sample Diluent. This reconstitution produces a stock solution of 4,000 pg/mL. Allow the standard to sit for about 10 minutes with gentle agitation prior to making serial dilutions (Making serial dilution in the wells directly is not permitted). Please firstly dilute the stock solution to 1,000 pg/mL and the diluted standard serves as the high standard (1,000 pg/mL). The Sample Diluent serves as the zero standard (0 pg/ml). pg/mL 4,000 1,000 500 250 125 62.5 31.2 15.6 0 Detection Reagent A and B - Dilute to the working concentration using Assay Diluent A or B (1:100), respectively.

Sample Collection: Plasma - Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples for 15 minutes at 1000 g at 2 - 8 within 30 minutes of collection. Store samples at -20 or -80 . Avoid repeated freeze-thaw cycles. Other biological fluids - Remove particulates by centrifugation and assay immediately or aliquot and store samples at -20 or -80 . Avoid repeated freeze-thaw cycles. Note: Plasma to be used within 7 days may be stored at 2-8, otherwise samples must stored at -20 ( 1 month) or -80 ( 2 months) to avoid loss of bioactivity and contamination. Avoid freeze-thaw cycles. When performing the assay slowly bring samples to room temperature.

Assay Procedure:

Allow all reagents to reach room temperature (Please do not dissolve the reagents at 37 directly.). All the reagents should be mixed thoroughly by gently swirling before pipetting. Avoid foaming. Keep appropriate numbers of strips for 1 experiment and remove extra strips from microtiter plate. Removed strips should be resealed and stored at 4 until the kits expiry date. Prepare all reagents, working standards and samples as directed in the previous sections. Please predict the concentration before assaying. If values for these are not within the range of the standard curve, users must determine the optimal sample dilutions for their particular experiments.
1. Add 100 of Standard, Blank, or Sample per well. Cover with the Plate sealer. Incubate for two hours at 37 .
2. Remove the liquid of each well, don’t wash.
3. Add 100 μl of Detection Reagent A working solution to each well. Cover with the Plate sealer. Incubate for 1 hour at 37 . Detection Reagent A working solution may appear cloudy. Warm to room temperature and mix gently until solution appears uniform.
4. Aspirate each well and wash, repeating the process three times for a total of three washes. Wash by filling each well with Wash Buffer (approximately 400 μl) using a 4 squirt bottle, multi-channel pipette, manifold dispenser or autowasher. Complete removal of liquid at each step is essential to good performance. After the last wash, remove any remaining Wash Buffer by aspirating or decanting. Invert the plate and blot it against clean paper towels.
5. Add 100 μl of Detection Reagent B working solution to each well. Cover with a new Plate sealer. Incubate for one hour at 37 .
6. Repeat the aspiration/wash process for five times as conducted in step
4. 7. Add 90 μl of Substrate Solution to each well. Cover with a new Plate sealer. Incubate for 15 - 30 minutes at 37 . Protect from light.
8. Add 50 μl of Stop Solution to each well. If color change does not appear uniform, gently tap the plate to ensure thorough mixing.
9. Determine the optical density of each well at once, using a microplate reader set to 450 nm.
Important Note:1. Absorbance is a function of the incubation time. Therefore, prior to starting the assay it is recommended that all reagents should be freshly prepared prior to use and all required strip-wells are secured in the microtiter frame. This will ensure equal elapsed time for each pipetting step, without interruption.
2. Please carefully reconstitute Standards or working Detection Reagent A and B according to the instruction, and avoid foaming and mix gently until the crystals have completely dissolved. The reconstituted Standards can be used only once. This assay requires pipetting of small volumes. To minimize imprecision caused by pipetting, ensure that pipettors are calibrated. It is recommended to suck more than 10μl for once pipetting.
3. To ensure accurate results, proper adhesion of plate sealers during incubation steps is necessary. Do not allow wells to sit uncovered for extended periods between incubation steps. Once reagents have been added to the well strips, DO NOT let the strips DRY at any time during the assay.
4. For each step in the procedure, total dispensing time for addition of reagents to the assay plate should not exceed 10 minutes.
5. To avoid cross-contamination, change pipette tips between additions of each standard level, between sample additions, and between reagent additions. Also, use separate reservoirs for each reagent.
6. The wash procedure is critical. Insufficient washing will result in poor precision and falsely elevated absorbance readings.
7. Duplication of all standards and specimens, although not required, is recommended.
8. Substrate Solution is easily contaminated. Please protect it from light.

Calculation of Results:

Average the duplicate readings for each standard, control, and sample and subtract the average zero standard optical density. Create a standard curve by reducing the data using computer software capable of generating a four parameter logistic (4-PL) curve-fit. As an alternative, construct a standard curve by plotting the mean absorbance for each standard on the x-axis against the concentration on the y-axis and draw a best fit curve through the points on the graph. The data may be linearized by plotting the log of the TF concentrations versus the log of the O.D. and the best fit line can be determined by regression analysis. It is recommended to use some related software to do this calculation, such as curve expert 13.0. This procedure will produce an adequate but less precise fit of the data. If samples have been diluted, the concentration read from the standard curve must be multiplied by the dilution factor.

Restrictions: For Research Use only

Handling

Handling Advice: 1. The kit should not be used beyond the expiration date on the kit label.
2. Do not mix or substitute reagents with those from other lots or sources.
3. If samples generate values higher than the highest standard, further dilute the samples and repeat the assay. Any variation in standard diluent, operator, pipetting technique, washing technique,incubation time or temperature, and kit age can cause variation in binding.
4. This assay is designed to eliminate interference by soluble receptors, ligands, binding proteins, and other factors present in biological samples. Until all factors have been tested in the Immunoassay, the possibility of interference cannot be excluded.
3.

Storage: 4 °C/-20 °C

Storage Comment: The Standard, Detection Reagent A, Detection Reagent B and the 96-well strip plate should be stored at -20 °C upon being received. The other reagents can be stored at 4 °C.

Target Details for FTH1

Target: FTH1 (Ferritin, Heavy Polypeptide 1 (FTH1))

Alternative Name: Fth1 (FTH1 Products)

Synonyms: FHC ELISA Kit, FTH ELISA Kit, FTHL6 ELISA Kit, PIG15 ELISA Kit, PLIF ELISA Kit, apoferritin ELISA Kit, ferritin ELISA Kit, fhc ELISA Kit, fth ELISA Kit, fth1 ELISA Kit, fthl6 ELISA Kit, ftn-2 ELISA Kit, pig15 ELISA Kit, plif ELISA Kit, fth1b ELISA Kit, Fth ELISA Kit, HFt ELISA Kit, MFH ELISA Kit, fb06g09 ELISA Kit, hm:zeh1145 ELISA Kit, wu:fb06g09 ELISA Kit, wu:fq18c10 ELISA Kit, zeh1145 ELISA Kit, ferritin heavy chain 1 ELISA Kit, ferritin, heavy polypeptide 1 L homeolog ELISA Kit, ferritin, heavy polypeptide 1 S homeolog ELISA Kit, ferritin heavy polypeptide 1 ELISA Kit, ferritin, heavy polypeptide 1a ELISA Kit, ferritin ELISA Kit, ferritin, heavy polypeptide 1 ELISA Kit, tudor domain containing 9 ELISA Kit, ferritin, heavy polypeptide 1 a ELISA Kit, FTH1 ELISA Kit, fth1.L ELISA Kit, fth1.S ELISA Kit, Fth1 ELISA Kit, fth1a ELISA Kit, EAMY_RS19690 ELISA Kit, fth1 ELISA Kit, TDRD9 ELISA Kit

Background: Ferritin is an iron-protein complex formed from an intracellular acceptor called Apoferritin. Apoferritin is a large molecular weight 450,000 protein produced by the liver. Iron as Fe (HO)3 linked to apoferritin is then stored in the cytoplasm of the reticuloendothelial system, liver, spleen and bone marrow. Ferritin is the body's iron storage protein functioning primarily as a site for iron storage from which iron may be mobilized in response to such stimuli a dietary change, blood loss or pregnancy. The direct quantitation of serum ferritin offers the physician a convenient and accurate measure of total body iron stores, by means of diagnosing iron-deficiency and anemia due to such causes as inflammation and hepatic or renal disease. In addition, serum ferritin concentration may be useful in detecting iron overload, which may allow the detection of idiopathic hemachromatosis in the precirrhotic storage.

Gene ID: 3164

Pathways: Transition Metal Ion Homeostasis