Rat IL18-BP ELISA Kit
SKU: 12189297328

Rat IL18-BP ELISA Kit

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Description

Rat IL18-BP ELISA KitProduct Specification Usage Experimental equipment required for the experiment: 1. Microplate reader (450nm) 2. High precision pipette and gun tips: 0. 5 10uL, 5 50uL, 20 200uL, 200 1000uL 3. 37 constant temperature box 4. Distilled water or deionized water Sample processing and requirements: 1. Serum: Place the whole blood sample collected in the serum separation tube at room temperature for 2 hours or at 4 overnight, then centrifuge at 1000g for 20

Product Specification

Usage Experimental equipment required for the experiment:
1. Microplate reader (450nm)
2. High-precision pipette and gun tips: 0.5-10uL, 5-50uL, 20-200uL, 200-1000uL
3. 37℃ constant temperature box
4. Distilled water or deionized water

Sample processing and requirements:
1. Serum: Place the whole blood sample collected in the serum separation tube at room temperature for 2 hours or at 4℃ overnight, then centrifuge at 1000×g for 20 minutes, and take the supernatant, or store the supernatant at -20℃ or -80℃, but avoid repeated freezing and thawing.
2. Plasma: Collect the specimen using EDTA or heparin as an anticoagulant. Centrifuge the specimen at 1000 × g for 15 minutes at 2-8°C within 30 minutes of collection. The supernatant can be assayed or stored at -20°C or -80°C, but avoid repeated freezing and thawing.
3. Tissue homogenization: Rinse the tissue with pre-chilled PBS (0.01M, pH 7.4) to remove residual blood (lysed red blood cells in the homogenate will affect the measurement results). Weigh the tissue and mince it. Add the minced tissue to the appropriate volume of PBS (generally a 1:9 weight-to-volume ratio, e.g., 1 g of tissue sample to 9 mL of PBS. The specific volume can be adjusted according to experimental needs and recorded. It is recommended to add protease inhibitors to the PBS) in a glass homogenizer and grind thoroughly on ice. To further lyse tissue cells, the homogenate can be sonicated or repeatedly frozen and thawed. Finally, centrifuge the homogenate at 5000 × g for 5-10 minutes, and the supernatant can be assayed.
4. Other biological fluids: Centrifuge at 1000xg for 20 minutes, remove the supernatant, and test.

Pre-test preparation:
1. Remove the test kit from the refrigerator 10 minutes in advance and equilibrate to room temperature.
2. Prepare the standard gradient working solution: Add 1 mL of universal diluent to the lyophilized standard, let it stand for 15 minutes to completely dissolve, then gently mix (concentration is 40 ng/mL). Then dilute to the following concentrations: 40 ng/mL, 20 ng/mL, 10 ng/mL, 5 ng/mL, 2.5 ng/mL, 1.25 ng/mL, 0.625 ng/mL, and 0 ng/mL. Serial dilution method: Take 7 EP tubes and add 500 μL of universal diluent to each tube. Pipette 500 μL of the 40 ng/mL standard working solution into the first EP tube and mix thoroughly to make a 20 ng/mL standard working solution. Repeat this procedure for subsequent tubes. The last tube serves directly as a blank well; there is no need to aspirate the liquid from the penultimate tube. See the figure below for details.

3. Preparation of Biotinylated Antibody Working Solution: 15 minutes before use, centrifuge the concentrated biotinylated antibody at 1000×g for 1 minute. Dilute the 100× concentrated biotinylated antibody to a 1× working concentration using universal diluent (e.g., 10µL concentrate + 990µL universal diluent). Prepare immediately before use.
4. Prepare the enzyme conjugate working solution: 15 minutes before use, centrifuge the 100× concentrated enzyme conjugate at 1000×g for 1 minute. Dilute the 100× concentrated HRP enzyme conjugate to a 1× working concentration with universal diluent (e.g., 10 μL of concentrate + 990 μL of universal diluent). Prepare immediately.
5. Prepare the 1× wash solution: Dispense 10 mL of 20× wash solution into 190 mL of distilled water (concentrated wash solution removed from the refrigerator may crystallize; this is normal. Allow to stand at room temperature until the crystals have completely dissolved before preparing).

Procedure:
1. Remove the desired strips from the aluminum foil bag after equilibration at room temperature for 10 minutes. Seal the remaining strips in a ziplock bag and return to 4°C.
2. Sample addition: Add 100 μL of sample or standard of varying concentrations to the corresponding wells. Add 100 μL of universal diluent to the blank wells. Cover with a film and incubate at 37°C for 60 minutes. (Recommendation: Dilute the sample to be tested at least 1-fold with universal diluent before adding it to the ELISA plate. This will reduce the impact of matrix effects on the test results. The sample concentration should be multiplied by the corresponding dilution factor when calculating the final sample concentration. It is recommended to run replicates for all test samples and standards.)
3. Add Biotinylated Antibody: Remove the ELISA plate and discard the liquid without washing. Add 100 μL of Biotinylated Antibody Working Solution directly to each well. Cover with a film and incubate at 37°C for 60 minutes.
4. Wash: Discard the liquid and add 300 μL of 1x Wash Solution to each well. Let stand for 1 minute, shake off the wash solution, and pat dry on absorbent paper. Repeat this process three times (a plate washer can also be used).
5. Add Enzyme Conjugate Working Solution: Add 100 μL of Enzyme Conjugate Working Solution to each well. Cover with a film and incubate at 37°C for 30 minutes.
6. Washing: Discard the liquid and wash the plate five times as in step 4.
7. Adding substrate: Add 90 μL of substrate (TMB) to each well, cover with a sealing film, and incubate at 37°C in the dark for 15 minutes.
8. Adding stop solution: Remove the ELISA plate and add 50 μL of stop solution directly to each well. Immediately measure the OD value of each well at a wavelength of 450 nm.

Calculating experimental results:
1. Calculate the average OD value of the standard and sample replicates and subtract the OD value of the blank well as a correction factor. Plot the standard curve of the four-parameter logistic function on double-logarithmic graph paper, with concentration as the horizontal axis and OD value as the vertical axis.
2. If the sample OD value is higher than the upper limit of the standard curve, dilute the sample appropriately and retest. Multiply the sample concentration by the corresponding dilution factor.

Theory This kit utilizes a double-antibody sandwich enzyme-linked immunosorbent assay (ELISA). Sample, standard, biotinylated detection antibody, and HRP conjugate are sequentially added to microwells pre-coated with Interleukin 18 Binding Protein (IL18-BP) capture antibody. After incubation and washing, the sample is developed using the substrate TMB. TMB is converted to blue by peroxidase (HRP) catalysis and to yellow by acid. The intensity of the color is positively correlated with the amount of Interleukin 18 Binding Protein (IL18-BP) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration.
Source Rat
Synonym Rat Interleukin 18 Binding Protein ELISA Kit
Detection Type Double antibody sandwich method
Composition
Name 9 6 T  match   set remark
Pre-coating 96 Well plate 8 Hole ×12 Strip without
Standard 2 branch
Dilute as per instructions
Universal diluent
2×20mL
without
Concentrated biotinylated antibody ( 100× )  
120uL
Dilute as per instructions
Concentrated enzyme conjugate ( 100× )
120uL
Dilute as per instructions
20× Washing liquid
2×10mL
Dilute as per instructions
Bottom thing ( TMB )
10mL
without
Stop liquid
6mL
without
Sealing film
4 Zhang
without
Instructions
1 Share
without
Background Interleukin-18 binding protein (IL18BP) is a protein encoded by the IL18BP gene. This gene encodes an inhibitor of the proinflammatory cytokine IL-18. This protein binds to IL-18, preventing it from binding to its receptor, thereby inhibiting IL-18-induced IFN-gamma production. This protein is constitutively expressed and secreted in monocytes. Its expression can be enhanced by IFN-gamma.
General Notes 1. Strictly adhere to the specified incubation time and temperature to ensure accurate results. All reagents must be at room temperature (20-25°C) before use. Refrigerate reagents immediately after use.
2. Improper plate washing may result in inaccurate results. Ensure that all liquid in the wells is aspirated thoroughly before adding substrate. Do not allow the wells to dry out during incubation.
3. Remove any residual liquid and fingerprints from the bottom of the plate, as this will affect the OD value.
4. The substrate developer solution should be colorless or very light in color. Do not use substrate solution that has turned blue.
5. Avoid cross-contamination of reagents and specimens to prevent erroneous results.
6. Avoid direct exposure to strong light during storage and incubation.
7. Do not expose any reagents to bleaching solvents or the strong fumes emitted by bleaching solvents. Any bleaching agent will destroy the biological activity of the reagents in the kit.
8. Do not use expired products, and do not mix components with different product numbers and batches.
9. Recombinant proteins from sources other than the kit may not be compatible with the antibodies in this kit and will not be recognized.
10. If there is a possibility of disease transmission, all samples should be managed properly and samples and testing devices should be handled according to prescribed procedures.
Storage Temp. If the unopened kit is stored at 4°C, the shelf life is 6 months.
Test Range 0.62-40 ng/mL
Applications Serum, plasma, tissue homogenates and other biological fluids
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SKU: 12189297328

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Clare Quilty
Boise, US
★★★★★ 5
A stark, brutish "Petulia" with a pistol in its pocket
It's about time this movie got released on DVD. It's odd that a film could spawn a remake ("Payback"), a glib nod ("Grosse Pointe Blank") and countless homages ("The Limey," among others) and still be as underseen as "Point Blank." The lack of a disc certainly didn't help its low profile, but of course this is a challenging, idiosyncratic movie, even three decades later. The plot is simple -- a crook is betrayed by his wife and partner and spends the rest of the movie trying to get what he's owed -- but the editing and narrative structure is unusual. What in the world did audiences possibly make of this back when it was first released? It's a remarkable film, as startling and innovative as Richard Lester's "Petulia," although admittedly it's thematically much less complex. This edition is excellent, too. Great sound, great picture and a fantastic commentary by director John Boorman and big-time "Point" fan Steven Soderbergh, who laughingly admits to Boorman that he's ripped this movie off more than a few times. Their chat is more technical than gossipy and deals heavily with the editing, the production (the script was only 70 pages long), the studio's concerns about the picture, the actors, violence, surrealism (is it all a dream?) and Boorman's elaborate use of color (the tones of clothing and sets intensify over the course of the film). I've gotten a lot of good DVD's this year but in terms of content, presentation and extas, this is one of the best.
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Reviewed in the United States on August 1, 2005
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Joe Movie
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★★★★★ 4
One of Marvins better offerings
Like the Killers before marvin was almost destined to play the part of Walker in this fast moving gut wrenching but always realistic thriller brilliantly collaborated by Marvin and Boorman who had no peer in this type of film.For those unenlightened souls who downplay Marvins career this was the one that to my mind surpassed most of his prior efforts with the exception of the Killers which was above par in all respects.Marvin was ahead of the pack in the 60s playing this type of hard nosed no nonsense gangster type, no other actor came close and type casted him to some extent in this type of role which unlike may actors became a positive in his career. Just to show his brilliance as an actor he gave us later comedy roles which produced more acting accolades than that material for which he was better known namely what we see in Point Blank.He carries the whole movie as did all the great actors of that era and many since which in itself is the hallmark of greatness. Marvin was a man who so perfectly personified the parts he was playing that often other actors looked wooden in comparison.He was one of the rare individuals who could take a small co starring role and end up being the star of the movie, no mean feat when you were up against the best in the business at that time and there were plenty in the 60 and 70s.To my mind Point Blank will always be a MARVIN film and this is not to downplay other good workmanlike performances in the film but it will always rank highly in Marvins body of work which is a cut above his contemporaries in the roles for which he was known and appreciated.As for the film it played out in fairly predictable fashion until the final scenes under the golden gate brige which gave a nice twist and left the viewer unsure whether Walker did in fact take his money or simply leave the scene of the set up empty handed. The obvious answer is that he waited till the coast was clear and took his money. It is hard to accept that he did otherwise.In retrospect a movie that stands up 40 odd years later and is just as watchable as it was in 67. No mean feat.
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Reviewed in the United States on May 20, 2010
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Patrick Selitrenny
Dallas, US
★★★★★ 5
A forerunner to Dirty Harry and Lee Marvin shining...
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