Human LIF ELISA Kit (A2990)

Name: Human LIF ELISA Kit
Brand: Antibodies.com
SKU: A2990
Price: 355.0 GBP
Availability: InStock

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Name

Human LIF ELISA Kit

Description

Human LIF ELISA Kit is a sandwich Enzyme-Linked Immunosorbent Assay (sELISA) designed for the in vitro quantitative determination of human LIF in serum, plasma, tissue homogenates, cell lysates, cell culture supernates or other biological fluids.

Assay Type

Sandwich (quantitative)

Principle of Assay

Human LIF ELISA Kit (A2990) employs the sandwich enzyme immunoassay technique for the quantitative measurement of human LIF in serum, plasma, tissue homogenates, cell lysates, cell culture supernates or other biological fluids. An antibody specific for LIF has been pre-coated onto a 96-well microtiter plate. The standards and test samples are added into the wells and the LIF present in each sample is bound to the wells by the immobilized antibody. Following incubation, the wells are washed and then incubated with Biotinylated Anti-LIF Antibody, which binds the captured LIF present in each well. HRP-Avidin conjugate is then added to the wells and the microtiter plate is incubated. Following incubation and washing, TMB substrate solution is then used to visualize the HRP enzymatic reaction by catalysis to produce a blue-coloured product that changes to yellow after addition of acidic stop solution. The density of yellow is proportional to the amount of LIF captured in each well. The concentration of LIF can then be calculated by reading the O.D. absorbance at 450nm in a microplate reader and referring to the standard curve.

Platform

Pre-coated Microplate (12 x 8 well strips)

Detection Type

Colourmetric

Instrument

Colorimetric Microplate Reader

Sample Type

Serum, plasma, tissue homogenates, cell lysates, cell culture supernates or other biological fluids.

Sensitivity

30.5 pg/ml

Product Range

78.1-5000 pg/ml

Assay Time

4h 30m

Reactivity

Human

Recovery

Sample Type	n	Range	Average
Serum	5	80% - 102%	91%
EDTA Plasma	5	81% - 100%	90%
Heparin Plasma	5	80% - 89%	84%

Linearity

Sample Type	1:2	1:4	1:8	1:16
Serum (n=5)	87-91%	87-107%	74-101%	92-97%
EDTA Plasma (n=5)	90-105%	84-101%	90-101%	79-108%
Heparin Plasma (n=5)	84-95%	92-105%	82-105%	89-91%

Precision

Intra-assay: CV < 10%, Inter-assay: CV < 12%

General Notes

Information online is representative. Always refer to the Product Manual inside the kit.

Storage

Store kit components at +4°C or -20°C as instructed. Do not use past expiration date!

Synonyms

D factor, Differentiation-stimulating factor, HILDA, Leukemia inhibitory factor, Melanoma-derived LPL inhibitor, MLPLI

Product Note

Information online is representative. Always refer to the Product Manual inside the kit.

Disclaimer

This product is for research use only. It is not intended for diagnostic or therapeutic use.

Kit Components

Item	Quantity	Storage
Pre-Coated 96 Well Microplate	12 x 8 Well Strips	-20°C
Lyopholized Standard	2 Vials	-20°C
Detection Solution A	120μl	-20°C
Detection Solution B	120µl	-20°C
Wash Buffer (30X)	20ml	+4°C
Sample Dilution Buffer	45ml	-20°C
TMB Substrate	9ml	+4°C
Stop Solution	6ml	+4°C
Plate Sealers	5 Adhesive Strips	-

Required Materials

The following materials are not included in the kit, but are required to run this assay:

Microplate reader capable of measuring absorbance at 450nm ± 10nm.
Squirt bottle, multichannel pipette reservoir, or automated microplate washer.
Graph paper or computer software capable of generating logarithmic functions.
Test tubes and Eppendorf tubes to prepare standards and samples.
Multi- and single-channel pipettes and sterile pipette tips.
Deionized or distilled water.
Absorbent paper towels.
37°C incubator.

Important Notes

Sample concentrations should be estimated before being used in the assay and a proper dilution factor should be selected to make the diluted target protein concentration fall in the optimal detection range of this kit.
Hemolysis can greatly impact the validity of test results. Take care to minimize hemolysis.
Samples should be brought to room temperature before starting the assay.
This ELISA kit is intended for the analysis of biological samples and has not been validated for purified/recombinant proteins, industrial, or commercial materials. Performance with such samples cannot be guaranteed, as differences in expression, sequence, tags, folding, post-translational modifications, purity, activity, or buffer composition may affect assay results. For further guidance, please contact our technical support team.

Bring all reagents and samples to room temperature before use.

We recommend that all standards and samples be assayed in duplicate.

Step 1

Prepare all reagents, samples, and working standards as directed in the previous sections.

Step 2

Determine the number of microplate strips required for the assay and insert them into the plate frame. Unused strips should be stored in a foil pouch at -20°C.

Step 3

Add 100μl of standard solutions to the standard wells. Note: A plate layout is provided to record standards and samples assayed.

Step 4

Add 100μl of samples to the sample wells.

Step 5

Cover with an adhesive plate sealer provided and incubate at 37°C for 2 hours.

Step 6

Remove the plate sealer and aspirate the liquid from the plate.

Step 7

Add 100μl of working Detection Solution A to each well.

Step 8

Cover with a new adhesive plate sealer and incubate at 37°C for 60 minutes.

Step 9

Remove the plate sealer, aspirate the liquid from the plate, and wash the plate three times with Wash Buffer. Soak wells with 300μl of Wash Buffer for 1-2 minutes each time. Note: 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. Blot the plate onto paper towels or other absorbent material. Do not let the wells dry completely at any time!

Step 10

Add 100μl of working Detection Solution B to each well.

Step 11

Cover with a new adhesive plate sealer and incubate at 37°C for 60 minutes.

Step 12

Remove the plate sealer, aspirate the liquid from the plate, and wash the plate five times with Wash Buffer. Soak wells with 300μl of Wash Buffer for 1-2 minutes each time.

Step 13

Add 90μl of TMB Substrate to each well. Cover with a new plate sealer and incubate at 37°C in the dark for 15-25 minutes (do not exceed 30 minutes). Note: This incubation time is for reference only, the optimal reaction time should be determined by the end-user and can be shortened or extended according to the actual color change. The reaction may be terminated when a gradient is apparent in the standard wells.

Step 14

Add 50μl of Stop Solution to each well. The color in the wells should change from blue to yellow immediately. Note: If the color in the wells is green or the color change does not appear uniform, gently tap the plate to ensure thorough mixing.

Step 15

Determine the optical density (O.D. value) of each well immediately using a microplate reader set to 450 nm.

Scientific Validation DataValidation Data (2)

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Standard Curve - Human LIF ELISA Kit (A2990)

Representative standard curve for Human LIF ELISA Kit (A2990).

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Human LIF ELISA Kit (A2990)

Tumor-derived LIF signaling mediates paracrine effects on muscle and hepatic cells. LIF concentrations in conditioned media from untreated (NT) and LIF-stimulated MKN45 spheroids were quantified by ELISA (top). Schematic illustration of the experimental workflow is shown below. MKN45 spheroids were either left untreated or stimulated with LIF, after which conditioned media were collected and used to treat C2C12 myoblasts and HepaRG hepatocytes.

Product Citations (1)

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(6)

Tumor-Derived LIF Promotes GDF15-Driven Cachexia and Adverse Outcomes in Gastric Cancer

This publication cites Human LIF ELISA Kit (A2990).

Article Abstract

Cancer cachexia is a multifactorial metabolic syndrome characterized by progressive skeletal muscle and adipose tissue loss, systemic inflammation, and poor clinical outcomes, and represents a major unmet clinical need in gastric cancer. Growth Differentiation Factor 15 (GDF15) is a key mediator of cachexia-associated anorexia and tissue wasting; however, the upstream mechanisms regulating its expression in gastric cancer remain poorly defined. Leukemia Inhibitory Factor (LIF), a pleiotropic cytokine implicated in tumor progression and metabolic dysregulation, has emerged as a potential regulator of cachexia-related pathways. Here, we investigated the association between LIF in regulating GDF15 expression and its relationship with metabolic, inflammatory, and body composition alterations in gastric cancer. Transcriptomic profiling of paired neoplastic and non-neoplastic gastric mucosa from 61 gastric cancer patients revealed a significant upregulation of both LIF and GDF15 in tumor tissue, with a strong positive correlation between their expression levels. High GDF15 expression was associated with reduced overall survival, a finding validated in independent TCGA-STAD and ACRG cohorts. Intratumoral bile acid profiling uncovered a marked enrichment of primary bile acids and a depletion of secondary bile acids, resulting in reduced levels of bile acids with endogenous LIF receptor (LIFR) antagonist activity; elevated primary, LIFR non-antagonist bile acids were associated with worse survival outcomes. Clinically, increased LIF and GDF15 expression correlated with weight loss, heightened inflammatory burden, reduced serum protein and albumin levels, and impaired body composition in a sub-cohort of 19 patients. Notably, LIF expression showed a significant inverse association with both lumbar skeletal muscle index (L3SMI) and subcutaneous adipose tissue index (SATI). Mechanistically, experimental models demonstrated that LIF enhances proliferative activity in gastric cancer spheroids and exerts paracrine effects that impair myogenic differentiation and suppress hepatic metabolic gene expression. Collectively, these findings identify the LIF/GDF15 axis as a central driver of cancer-associated cachexia in gastric cancer and highlight LIF signaling as a potential therapeutic target.

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