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Description
Rat PKCd ELISA KitProduct Specification Usage Required experimental equipment: 1. Microplate reader (450nm) 2. High precision pipettes and pipette tips: 0. 5 10uL, 5 50uL, 20 200uL, 200 1000uL 3. 37C incubator 4. Distilled or deionized water Sample preparation and requirements: 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 and
Product Specification
| Usage | Required experimental equipment: 1. Microplate reader (450nm) 2. High-precision pipettes and pipette tips: 0.5-10uL, 5-50uL, 20-200uL, 200-1000uL 3. 37°C incubator 4. Distilled or deionized water Sample preparation and requirements: 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 and mince the tissue. Add the minced tissue to the appropriate volume of PBS (generally a 1:9 weight-to-volume ratio, e.g., 1g of tissue sample to 9mL 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 collect the supernatant for analysis. Cell Lysis Buffer: Adherent cells should be gently washed with pre-chilled PBS, then trypsinized and harvested by centrifugation at 1000×g for 5 minutes. Suspension cells can be harvested directly by centrifugation. Collected cells should be washed three times with pre-chilled PBS and resuspended in 150-200 μL of PBS per 1×10^6 cells (it is recommended to add protease inhibitors to the PBS; if the cell count is very low, reduce the PBS volume appropriately). Disrupt the cells by repeated freezing and thawing or sonication. Centrifuge the extract at 1500×g for 10 minutes at 2-8°C, and collect the supernatant for analysis. 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 10 ng/mL). Then dilute to the following concentrations: 10 ng/mL, 5 ng/mL, 2.5 ng/mL, 1.25 ng/mL, 0.625 ng/mL, 0.3125 ng/mL, 0.15625 ng/mL, and 0 ng/mL. Serial dilution method: Take seven EP tubes and add 500uL of universal diluent to each. Pipette 500uL of the 10ng/mL standard working solution into the first EP tube and mix thoroughly to make a 5ng/mL standard working solution. Repeat this procedure for subsequent tubes. The last tube serves as a blank well; there is no need to pipette liquid from the penultimate tube. See the figure below for details. 3. Preparation of biotinylated detection antibody working solution: Centrifuge the concentrated biotinylated antibody at 1000×g for 1 minute 15 minutes before use. Dilute the 100× concentrated biotinylated antibody to a 1× working concentration with universal diluent (e.g., 10uL concentrate + 990uL universal diluent). Prepare and use immediately. 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. |
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| Theory | This kit uses a double-antibody sandwich enzyme-linked immunosorbent assay (ELISA). Sample, standard, biotin-labeled detection antibody, and HRP conjugate are sequentially added to microwells pre-coated with a capture antibody against Protein Kinase C Delta type (PKCd). After incubation and washing, the sample is developed using the substrate TMB. TMB is converted to blue by HRP peroxidase and to yellow by acid. The intensity of the color is positively correlated with the amount of Protein Kinase C Delta type (PKCd) in the sample. The absorbance (OD) is measured at 450 nm using a microplate reader to calculate the sample concentration. | |||||||||||||||||||||||||||||||||
| Source | Rat | |||||||||||||||||||||||||||||||||
| Synonym | Rat Protein Kinase C Delta type ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Double antibody sandwich method | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | Protein kinase C delta (PKCδ), also known as PRCKD or NPKC-delta, is an enzyme encoded by the PRKCD gene. The protein encoded by this gene is a member of the protein kinase C (PKC) family. PKCs are a family of serine- and threonine-specific protein kinases that are activated by the second messenger diacylglycerol. PKC family members phosphorylate a variety of protein targets and are known to participate in numerous cellular signaling pathways. PKC family members also serve as primary receptors for phorbol esters, a class of tumor promoters. | |||||||||||||||||||||||||||||||||
| 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. |
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| Storage Temp. | If the unopened kit is stored at 4°C, the shelf life is 6 months. | |||||||||||||||||||||||||||||||||
| Test Range | 0.156-10 ng/mL | |||||||||||||||||||||||||||||||||
| Applications | Tissue homogenates, cell lysates, and other biological fluids |
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4.6 ★★★★★
Based on 22 reviews
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Product Reviews
★★★★★ 5
Worth viewing and studying.
First major Kubrick feature film noir. Nothing about Sterling Hayden's Communism here. Cool caper with a classic cast. The flick provides an impactful reminder to never skimp on low quality luggage, to use a double lock, and to take time whenever possible to bundle your cash with rubber bands.
"While playing chess in Washington Square, Kubrick met producer James B. Harris, who considered Kubrick to be "the most intelligent, most creative person I have ever come in contact with", and the two formed the Harris-Kubrick Pictures Corporation in 1955.[52] Harris purchased the rights to Lionel White's novel Clean Break for $10,000,[g] and upon Kubrick's suggestion, they hired film noir novelist Jim Thompson to write the script for the film—which later became The Killing (1956)—about a meticulously planned racetrack robbery gone wrong. The film starred Sterling Hayden, with whom Kubrick had been impressed in The Asphalt Jungle (1950).[54] Kubrick and Harris moved to Los Angeles from New York and signed with the Jaffe Agency to shoot the picture, which became Kubrick's first full-length feature film shot with a professional cast and crew. The Union in Hollywood stated that Kubrick would not be permitted to be both the director and the cinematographer of the movie, so veteran cinematographer Lucien Ballard was hired for the shooting. Kubrick agreed to waive his fee for the production, which was shot in just 24 days on a budget of $330,000.[55] He clashed with Ballard during the shooting, and on one occasion Kubrick threatened to fire Ballard following a camera dispute, despite being only 27 years old at the time and 20 years Ballard's junior.[54] Hayden recalled that Kubrick was "cold and detached. Very mechanical, always confident. I've worked with few directors who are that good".[56] The Killing failed to secure a proper release across the United States; the film made little money, and was promoted only at the last minute, as a second feature to the Western movie Bandido! (1956). Several contemporary critics lauded the film, however, with a reviewer for TIME comparing its camerawork to that of Orson Welles.[57] Today, critics generally consider The Killing to be among the best films of Kubrick's early career; its nonlinear narrative and clinical execution also had a major influence on later directors of crime films, including Quentin Tarantino. Dore Schary of Metro-Goldwyn-Mayer was highly impressed as well, and offered Kubrick and Harris $75,000 to write, direct, and produce a film, which ultimately became Paths of Glory (1957)." [Wikipedia]
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Reviewed in the United States on February 21, 2016
★★★★★ 5
Quintessential 50's Kubrick Noir!
Criterion continues to keep them coming. The Killing(1956), Stanley Kubrick's great noir adds to the recent influx of Criterion's recent titles in 2011, which closely follows Kiss Me Deadly(1955). What a feast for Noir addicts! Now we have another pristine upgrade of a print which greatly improves past releases in both quality as well as great special features. This time we get two jam filled discs of a very important Noir. This two-dvd special edition includes a bounty of goodies for lovers of Noir. There is a new digital restoration, which is excellent, as well as a new interview with producer James B Harris. Interviews with lead actor Sterling Haden, who is excellent in his role as the small-time criminal who plans a dangerous race-track heist with help from a corrupt cop, and an interview with author Robert Polito about writer Jim Thompson. That is just on the first disc, Disc two includes a richly restored high-def digital transfer of Kubrick's 1955 feature Killer's Kiss, new video appreciation of Killer's Kiss by film critic Geoffrey O'Brien, and a new trailer. Of course, you also get a 20 page booklet featuring an essay by film historian Haden Guest and an interview with actress Marie Windsor, which is a reprint but still quite good. Now on to the feature. The Killing was Kubrick's 3rd feature, and to most film historians, the one that put him on the map, although some people would favor Paths Of Glory which was released in the next year 1957 as his breakthrough as a major director. I beg to differ. The Killing is told in a non-linear style which many movie goers have difficulty following, even now in the 21st Century. But to lovers of Noir, by 1956 they had become quite used to it and had no problem with it. In fact, many noir lovers enjoyed putting the pieces together which to them, only added to the experience. The film displays what has become a very familiar Kubrick theme. That is the breakdown, malfunction or fallibility of man and his plans. Just as in Kubrick's subsequent films such as Dr. Strangelove and further on to 2001:A Space Odyssey which became major mainstream successes. His manipulation of time in bits and pieces differs most strikingly from 40's Noir, such as The Killers and Out Of The Past. As players inthis game are established, the film leaps backwards until all of the parts fit together like the pieces of a jigsaw puzzle. The well planned heist falls apart just like the suitcase full of money falls off a cart and scatters the bills in the wind. Just as they say, "The Best Laid Plans". In addition to Hayden, the other members of the heist, especially Vince Edwards, Elisha Cook Jr, and especially Timothy Carey, are all excellent. Most definitely, pay the extra money and get this edition. SMRZ!!!
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Reviewed in the United States on September 15, 2011
★★★★★ 4
Worth Watching Crime Drama
I almost stopped watching this film early on because I found the behavior of the cashier's wife so annoying and insufferable. I stuck with it though. It is almost as if it is a crime re-enactment show because of the voiceover narration. In one of the close to final scenes, I thought "This is Kubrick, alright." It is interesting that I watched a film yesterday, "Dead End" in which some of the characters believe that the only life worth living is one with material wealth as do some of the characters in this film. How sad. Maybe that belief is what kills what has real worth. Maybe it is that belief that leads one to a dead end.
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Reviewed in the United States on October 6, 2025
★★★★★ 5
Excellent.
kubrick's first i think. an excellent b/w noir crime film. very influential. sets the standard. must see.
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Reviewed in the United States on June 5, 2026
★★★★★ 5
2 Early Kubrick Films in 1 Thrilling Package
The most important thing for the buyer to know about The Killing Criterion Blu-Ray is that it collects 2 of Stanley Kubrick’s earlier films: The Killing (1956) and Killer’s Kiss (1955). Even though the Killer’s Kiss title is not on the box, it is present as an extra, along with its trailer.
Other than that, you can expect the usual from a Criterion Collection release. For those who don’t know, that means terrific quality and presentation, with loads of extra supplementary material.
As far as the movies themselves go — unlike Fear and Desire, I can actually recommend these two movies as movies, as opposed to morbid curiosities of Kubrick fanatics. They are both pretty intense crime thrillers and just a ton of fun. This is Kubrick before he makes big studio movies with big budgets, but after he’s gained more of the experience and sophistication he would later be known for. The product feels raw, gritty, sometimes intimate, and gives you a look at his NYC roots. Kubrick is still an underdog at this stage in his career. So, would definitely recommend for Kubrick fans, and just crime thriller fans in general
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Reviewed in the United States on December 11, 2022
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