Technical guide & protocol Tech Support

Product Specifications

Analytical sensitivity

<0.01 pg/mL

Assay range

0.0128 - 5000 pg/mL

Sample type/volume

Serum
2-5 µL
Supernatant
2-5 µL
Plasma
2-5 µL

Hands-on time

30 min

Time-to-result

2 hr

Homogenous (no wash)

Yes

Instrument

qPCR Instruments

Product size

96 Tests

Contents

Assay Dilution Buffer
Antibody-Conjugate A
Antibody-Conjugate B
Antibody-Conjugate Dilution Buffer
Standard
Master Mix
Ligase

Shipping conditions

Dry ice

Storage

-20°C

Protein name

TNF alpha

Species (tested)

Human

Assay kit format

Proximity-based Immunoassay

Label or dye

FAM™

About This Kit

The ProQuantum Human TNF alpha Immunoassay Kit is designed to provide highly sensitive quantitative measurements of human TNF alpha protein in small sample volumes. Utilizing a proximity-based amplification technology, the assay combines the analyte specificity of high-affinity antibody-antigen binding with the signal detection and amplification capabilities of real-time PCR to achieve a simple yet powerful next-generation protein quantitation platform. A user-friendly workflow combined with intuitive Cloud-based software for analytics enables sample-to-answer in just 2 hours.

• High sensitivity-detect low levels of protein with greater sensitivity than traditional methods

• Broad dynamic range-≥5 logarithmic units, minimizing sample dilutions to ensure they fall within the range

• Small sample consumption-use 2-5 µL of sample (compared to 75 µL for triplicate wells with other methods)

• Fast, easy workflow-2 hours from sample to answer, with no wash steps

• No proprietary instrument to purchase-runs on any real-time PCR instrument

ProQuantum immunoassays utilize a matched pair of target-specific antibodies, each conjugated to a DNA oligonucleotide. During antibody-analyte binding, the two DNA oligos are brought into close proximity, which allows for ligation of the two strands and subsequent creation of a template strand for amplification. This platform leverages the sensitivity and large dynamic range of Applied Biosystems TaqMan real-time PCR technology (figure below).

The assay workflow is fast and easy-2 steps in 2 hours. There are a total of 7 components in each kit (figure below). First, mix the antibody-conjugates, dilute the sample, and create the standard curve in a working plate. Then, using a multi-channel pipette, add the antibody-conjugates and sample (or standard) into the wells of a PCR plate and incubate for 1 hour. Combine the master mix and ligase and add to the wells of the PCR plate, then run the plate on any qPCR instrument. After the run is complete, import the results file into the ProQuantum cloud-based software at https://apps.thermofisher.com/apps/proquantum. Using this software, the data can be analyzed easily to obtain protein concentration values. The software allows you to set up standard curves, design plate layouts, set up customized assay instructions, and obtain robust statistical group-wise comparisons.

Target information

TNF alpha is a multifunctional proinflammatory cytokine that belongs to the tumor necrosis factor (TNF) superfamily. This cytokine is mainly secreted by macrophage and bind to its receptors, TNFRSF1A/TNFR1 and TNFRSF1B/TNFBR. TNF alpha is involved in the regulation of immune cells, cell proliferation, differentiation, apoptosis, lipid metabolism, and coagulation. TNF alpha exists as a multimer of two, three, or five noncovalently linked units, but shows a single 17 kDa band following SDS-PAGE under non-reducing conditions. Knockout studies in mice also suggested the neuroprotective function of TNF alpha, and has been observed to causes tumor necrosis when injected into tumor-bearing mice. Other functions of TNF-alpha include its role in the immune response to bacterial, viral, parasitic and certain fungal infections, as well as its role in the necrosis of specific tumors. TNF alpha causes cytolysis or cytostasis of certain transformed cells, being synergistic with interferon-gamma in its cytotoxicity. This cytokine has been implicated in a variety of diseases, including autoimmune diseases, insulin resistance, and cancer.

For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.

Bioinformatics

Gene aliases : DIF, TNF, TNF-alpha, TNFA, TNFSF2, TNLG1F

Gene ID : (Human) 7124

Gene symbol : TNF

Protein Aliases : APC1 protein, Cachectin, TNF, macrophage-derived, monocyte-derived, TNF-a, TNF-alpha, Tumor necrosis factor, tumor necrosis factor ligand 1F, Tumor necrosis factor ligand superfamily member 2, tumor necrosis factor-alpha

UniProt ID (Human) P01375

guarantee_icon

Performance Guarantee

If an Invitrogen™ antibody doesn't perform as described on our website or datasheet,we'll replace the product at no cost to you, or provide you with a credit for a future purchase.*

Learn more
help_icon

We're here to help

Get expert recommendations for common problems or connect directly with an on staff expert for technical assistance related to applications, equipment and general product use.

Contact tech support