Huabio provides two different types of ELISA kits to meet your diverse needs

Quality ELISA Kit

Contains all components for detecting this protein target. (Different kits may vary slightly, but generally include detection antibody, recombinant standard, horseradish peroxidase-conjugated streptavidin, antibody-coated 96-well microplate, sealing film, chromogenic solution, stop solution, coating buffer (PBS), washing buffer, etc.)

DTSet Auxiliary Kit:

Contains capture antibody, detection antibody, recombinant standard, horseradish peroxidase-conjugated streptavidin (Streptavidin-HRP)

Others can be purchased separately:

96-well microplate, sealing film, chromogenic solution, stop solution, coating buffer (PBS), washing buffer, and detection buffer.

Huabio kits can be stored at 2-8 ℃ for 6 months after receipt.

The sensitivity of ELISA kits is usually expressed as the Limit of Detection (LOD) or Limit of Quantification (LOQ), which refers to the lowest analyte concentration that can be significantly distinguished from the blank background in a statistical sense. This value can be calculated by repeatedly measuring blank samples and low-concentration standards based on their signal values and standard deviations. Specific sensitivity data can be found in the product manual or detailed parameter table on the official website. Different kits have different sensitivity performance due to factors such as antibody affinity, enzyme labeling efficiency, and detection system.

If extremely weak or no color development is observed for the standard in the experiment, it may involve problems in multiple links, including but not limited to:

Improper reagent storage conditions：Such as enzyme conjugates or substrate solutions not being stored in the dark or being repeatedly frozen and thawed, resulting in decreased activity;

Insufficient incubation time or temperature：Failure to follow the instructions for incubation;

Decreased antibody titer：Capture antibodies or detection antibodies may have decreased activity due to improper storage or repeated use;

Insufficient washing steps：Residual substances interfere with the binding reaction;

Incorrect standard preparation：Incorrect selection of dilution buffer or gradient dilution operation.

It is recommended to systematically check the experimental process and ensure that each step of operation is consistent with the kit requirements. If the cause cannot be determined, please contact our technical support team and provide detailed experimental records for further analysis.

High background usually stems from non-specific binding or contamination, mainly including:

Insufficient washing：Failure to thoroughly remove unbound antibodies or enzyme conjugates, should ensure that the washing solution is fully separated and patted dry;

Reagent contamination or cross-contamination：Such as contamination of pipettes, troughs, or reagents by microorganisms or substances from previous experiments;

Sample matrix effects：Presence of endogenous enzymes or heterophilic antibodies in serum, plasma, or cell culture medium;

Insufficient blocking：Insufficient blocking solution concentration or incubation time, failing to effectively cover non-specific binding sites;

Excessive antibody concentration：Especially when the detection antibody usage exceeds the recommended range.

Optimizing the washing procedure, ensuring reagent cleanliness, appropriately diluting samples, or re-optimizing blocking conditions can help reduce background.

We strongly recommend using freshly prepared samples for testing to avoid degradation or denaturation. If storage is necessary, please follow these principles:

Short-term storage：Store in 4°C refrigeration, recommended to use within 3 days;

Long-term storage：Aliquot and store at –20°C or –80°C, avoid repeated freeze-thaw cycles (generally no more than 3 freeze-thaw cycles);

Note sample type differences：Some cytokines or phosphorylated proteins are easily degraded and require protease inhibitors and preferential storage at –80°C.

Please refer to the relevant recommendations in the kit manual for specific sample type storage requirements.

To ensure the reliability and repeatability of experimental results, we recommend:

Set duplicate wells：Each sample or standard should have at least 2 duplicate wells (in duplicate), and triplicate wells can further improve data reliability;

Set quality control：Including blank wells, negative controls, and positive controls;

Standard curve fitting：Use appropriate mathematical models (such as four-parameter logistic regression) to fit standard curves, and ensure R² > 0.99;

Data verification：Calculate the coefficient of variation (CV) between duplicate wells, usually requiring CV < 10%.

In addition, it is recommended to conduct pre-experiments to determine the optimal dilution multiple of samples.