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Prostate Specific Antigen (Total, human) ELISA Kit

Sensitive, Colorimetric Assay
Catalog #: K7431
$685.00

Product Details

Cat # +Size K7431-100
Size 100 assays
Detection Method Absorbance (450 nm)
Species Reactivity human
Applications This ELISA kit is used for quantitative detection of total Prostate Specific Antigen (PSA)
Features & Benefits • Easy, convenient and time-saving method to measure total Prostate Specific Antigen (PSA)
• Sensitivity: 0.0144 ng/ml
• Cross Reactivity: No significant cross-reactivity or interference between this analyte and its analogues was observed.
Kit Components • Microwells coated with Streptavidin
• PSA Standard*
• Anti-PSA Conjugate Reagent
• Wash Buffer (20X)
• TMB Substrate
• Stop Solution
Storage Conditions 2-8°C
Shipping Conditions Gel Pack
USAGE For Research Use Only! Not For Use in Humans.

Details

Prostate Specific Antigen (PSA) is useful in the management of patients with prostate cancer. It is a single chain glycoprotein produced by epithelial cells of the prostate gland. The measurement of serum PSA has become the most accepted test to indicate men who are at risk of having prostate cancer and who should be examined by other tests. Using a cut-off of 4 ng/ml, 92% of men over 50 years of age with malignant prostatic tissues, 8% of healthy men and 28% of men with benign prostate hyperplasia (BPH) test positive for PSA. Three major forms of PSA exist in the serum: free PSA, bound PSA and complex PSA. Bound PSA is found in higher concentrations in patients with prostate cancer; whereas, free PSA is detected in higher concentrations in patients with BPH. If the free PSA to total PSA ratio is >25%, it is unlikely that the patient has prostate cancer; whereas, if free PSA is <16% then prostate cancer is likely to be the cause. Serial measurement of PSA concentration in the serum is an important tool in monitoring patients with prostatic cancer and determining the potential and actual effectiveness of surgery or other therapies, or may allow for earlier discovery of residual or recurrent carcinoma after radical prostatectomy or radiotherapy. BioVision’s PSA ELISA kit is a solid phase assay based on a streptavidin-biotin principle. The standards, samples and a reagent mixture of Anti-PSA Enzyme and Biotin conjugates (conjugate reagent) are added into the wells, coated with Streptavidin. PSA in the serum sample forms a sandwich between two highly specific Anti-PSA antibodies, labeled with Biotin and HRP. Simultaneously, the biotinylated antibody is immobilized onto the well through a high affinity Streptavidin-Biotin interaction. Unbound protein and excess biotin/enzyme conjugated reagent are washed off, by washing buffer. Upon the addition of the substrate, the intensity of color developed is directly proportional to the concentration of PSA in the samples. A standard curve is prepared relating color intensity to the concentration of the PSA.


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What is a sandwich ELISA?
The analyte of interest in a sample is "sandwiched" by an immobilized capture antibody coated on the plate and a labeled antibody used for detection. Sandwich ELISA is very sensitive. The density of color is proportional to the amount of analyte captured from the samples and can be quantified when compared with standard curve.
What is a competitive ELISA?
The endogenous unlabled analyte of interest in a sample is "competed" by the exogenous labeled antigen coated on the plate for a limited amount of antibody binding sites. Therefore, the lower signal indicate higher concentration of the analyte.
What dilution range should I use for the samples?
A preliminary experiment is always recommended when working with new samples and ELISA Kits to ensure all results fall within the detection range.
Can I extend the standard curve (in either direction)?
Extended standard curve is not recommended by BioVision.
What type of software is needed to graph a 4-parameter or 5-parameter curve?
SoftMax Pro by Molecular Devices, SigmaPlot® by Systat Software Inc., or others can be used for this purpose.
Low absorbance; No signal
Target present below detection limits of assay
Reduce dilution factor to increase sample concentration  (Preliminary experiment is recommended for optimal dilution range)
Incompatible sample types
Use samples recommended by the kit
Standard dilutions are unstable
Use fresh standard dilutions
Buffers/substrates are contaminated
Use new buffers and substrates
Insufficient incubation time 
Extend incubation time
Cover or seal plates during all incubation 
Plate washings too vigorous
Pipette wash buffer gently or make sure the automatic wash system has correct pressure
Wells scratched with pipette or washing tips
Use caution when dispensing and aspirating into and out of wells
Incubation temperature too low
Ensure incubations are carried out at temperatures recommended by the manual
Reagents are cold
Bring all reagents to room temperature before each assay
Plate read at incorrect wavelength

Ensure the plate reader is set at the correct wavelength recommended on the protocol

High Background
Insufficient washing
Increase number of washes; Add a 30 second soaking step in between washes;  at the end of each washing step, invert plate on absorbent tissue to completely drain all wells
Residual buffer/substrates remain in wells
Remove all residual buffer and substrates at the end of each wash
Too much HRP-Streptavidin
Centrifuge and mix the substrate vial before use to avoid percipitation and inconsistent HRP concentration
Excessive incubation time
Reduce incubation time
Substrate incubation carry out in light or wait too long to read plate after adding stop solution
Avoid light during incubation and read signals immediately after adding stop solution
Standards improperly reconstituted or diluted
Briefly spin vials before opening; inspect for undissolved material after reconstitution
Poor standard curve
Standard improperly diluted
Confirm dilutions are made correctly
Standard dilutions are unstable
Use freshly diluted standards
Pipetting error 
use calibrated pipette and stay consistent between each pipette
Curve doesn't fit scale
Try plotting using different scales  (log-log, 4 parameter logistic, 5 parameter etc.)
Large intra-coefficient of variation (CV)
Bubbles in wells
Avoid bubbles during experiment, eliminate all bubbles prior to reading
Insufficient washing
Increase number of washes; add a 30 second soak step in between washes;  at the end of each washing step, invert plate on absorbent tissue to completely drain all wells
Wells not washed equally/thoroughly
Use calibrated (automated) multi-channel pipette for consistent handling, make sure all parts of the automated washer are unobstructed
Edge effects
Seal the plate completely during incubation
Incomplete reagent mixing
Make sure all reagents are mixed thoroughly during each step
Inconsistent sample preparation or storage
Consistent sample preparation. Optimize sample storage conditions and minimize freeze and thaw cycles
Stacked plates
Avoid stacking plates during incubation
Plate sealers not used or reused
Cover assay plate with plate sealer, use a fresh sealer each time to prevent contamination
Large inter-coefficient of variation (CV))
Insufficient washing
Increase number of washes; add a 30 second soak step in between washes;  at the end of each washing step, invert plate on absorbent tissue to completely drain all wells
Inconsistent incubation temperature
Make sure to follow recommended incubation temperatures. Avoid fluctuations in temperature due to environmental conditions.
Plate sealers not used or reused
Cover assay plate with plate sealer, use a fresh sealer each time to prevent contamination