Description
Human Lactotransferrin (LTF) ELISA Kit
The Human Lactotransferrin (LTF) ELISA Kit is a reliable and accurate tool for detecting lactotransferrin levels in human biological samples such as serum, plasma, and cell culture supernatants. This kit promises high sensitivity and specificity, ensuring consistent and precise results for various research applications.Lactotransferrin, also known as lactoferrin, is a multifunctional protein with antimicrobial, immunomodulatory, and iron-binding properties. It plays a crucial role in various biological processes, including immune defense, iron metabolism, and tissue repair.
Dysregulation of lactotransferrin levels has been implicated in conditions such as inflammation, infection, and cancer, making it a valuable biomarker for studying these diseases and exploring potential therapeutic interventions.Overall, the Human Lactotransferrin (LTF) ELISA Kit is an essential tool for researchers looking to investigate the role of lactotransferrin in health and disease, providing accurate and reliable results for their experiments.
| Product Name: | Human Lactotransferrin (LTF) ELISA Kit |
| SKU: | HUEB0854 |
| Size: | 96T |
| Target: | Human Lactotransferrin (LTF) |
| Synonyms: | Growth-inhibiting protein 12, Talalactoferrin, Lactoferrin, GIG12, LF |
| Assay Type: | Sandwich |
| Detection Method: | ELISA |
| Reactivity: | Human |
| Detection Range: | 0.78-50ng/mL |
| Sensitivity: | 0.052ng/mL |
| Intra CV: | 4.3% | ||||||||||||||||||||
| Inter CV: | 7.5% | ||||||||||||||||||||
| Linearity: |
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| Recovery: |
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| Function: | Isoform DeltaLf: transcription factor with antiproliferative properties and ability to induce cell cycle arrest. Binds to the DeltaLf response element found in the SKP1, BAX, DCPS, and SELENOH promoters. |
| Uniprot: | P02788 |
| Sample Type: | Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids |
| Specificity: | Natural and recombinant human Lactotransferrin |
| Sub Unit: | Monomer. Found in a complex with LTF, CLU, EPPIN and SEMG1. |
| Research Area: | Cancer |
| Subcellular Location: | Isoform DeltaLf Cytoplasm Nucleus Mainly localized in the cytoplasm. |
| Storage: | Please see kit components below for exact storage details |
| Note: | For research use only |
| UniProt Protein Function: | LTF: Transferrins are iron binding transport proteins which can bind two Fe(3+) ions in association with the binding of an anion, usually bicarbonate. Belongs to the transferrin family. 2 isoforms of the human protein are produced by alternative promoter. |
| UniProt Protein Details: | Protein type:Secreted; Protease; EC 3.4.21.-; Secreted, signal peptide Chromosomal Location of Human Ortholog: 3p21.31 Cellular Component: cell surface; cytoplasm; extracellular region; extracellular space; nucleus; protein complex; secretory granule; specific granule Molecular Function:heparin binding; iron ion binding; protein binding; protein serine/threonine kinase activator activity Biological Process: activation of NF-kappaB transcription factor; antibacterial humoral response; antifungal humoral response; cell redox homeostasis; cellular protein metabolic process; humoral immune response; innate immune response in mucosa; iron assimilation by chelation and transport; negative regulation of apoptosis; negative regulation of ATPase activity; negative regulation of lipopolysaccharide-mediated signaling pathway; negative regulation of viral genome replication; negative regulation of viral reproduction; positive regulation of I-kappaB kinase/NF-kappaB cascade; positive regulation of osteoblast differentiation; positive regulation of osteoblast proliferation; positive regulation of toll-like receptor 4 signaling pathway; regulation of cytokine production; regulation of tumor necrosis factor production; retinal homeostasis |
| NCBI Summary: | This gene is a member of the transferrin family of genes and its protein product is found in the secondary granules of neutrophils. The protein is a major iron-binding protein in milk and body secretions with an antimicrobial activity, making it an important component of the non-specific immune system. The protein demonstrates a broad spectrum of properties, including regulation of iron homeostasis, host defense against a broad range of microbial infections, anti-inflammatory activity, regulation of cellular growth and differentiation and protection against cancer development and metastasis. Antimicrobial, antiviral, antifungal and antiparasitic activity has been found for this protein and its peptides. Alternatively spliced transcript variants encoding different isoforms have been found for this gene. [provided by RefSeq, Sep 2014] |
| UniProt Code: | P02788 |
| NCBI GenInfo Identifier: | 85700158 |
| NCBI Gene ID: | 4057 |
| NCBI Accession: | P02788.6 |
| UniProt Secondary Accession: | P02788,O00756, Q16780, Q16785, Q16786, Q16789, Q5DSM0 Q8IU92, A8K9U8, B2MV13, B7Z4X2, E7EQH5, |
| UniProt Related Accession: | P02788 |
| Molecular Weight: | 73,161 Da |
| NCBI Full Name: | Lactotransferrin |
| NCBI Synonym Full Names: | lactotransferrin |
| NCBI Official Symbol: | LTF |
| NCBI Official Synonym Symbols: | LF; HLF2; GIG12; HEL110 |
| NCBI Protein Information: | lactotransferrin |
| UniProt Protein Name: | Lactotransferrin |
| UniProt Synonym Protein Names: | Growth-inhibiting protein 12; Talalactoferrin |
| Protein Family: | Lactotransferrin |
| UniProt Gene Name: | LTF |
| UniProt Entry Name: | TRFL_HUMAN |
| Component | Quantity (96 Assays) | Storage |
| ELISA Microplate (Dismountable) | 8×12 strips | -20°C |
| Lyophilized Standard | 2 | -20°C |
| Sample Diluent | 20ml | -20°C |
| Assay Diluent A | 10mL | -20°C |
| Assay Diluent B | 10mL | -20°C |
| Detection Reagent A | 120µL | -20°C |
| Detection Reagent B | 120µL | -20°C |
| Wash Buffer | 30mL | 4°C |
| Substrate | 10mL | 4°C |
| Stop Solution | 10mL | 4°C |
| Plate Sealer | 5 | - |
Other materials and equipment required:
- Microplate reader with 450 nm wavelength filter
- Multichannel Pipette, Pipette, microcentrifuge tubes and disposable pipette tips
- Incubator
- Deionized or distilled water
- Absorbent paper
- Buffer resevoir
*Note: The below protocol is a sample protocol. Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.
Allow all reagents to reach room temperature (Please do not dissolve the reagents at 37°C directly). All the reagents should be mixed thoroughly by gently swirling before pipetting. Avoid foaming. Keep appropriate numbers of strips for 1 experiment and remove extra strips from microtiter plate. Removed strips should be resealed and stored at -20°C until the kits expiry date. Prepare all reagents, working standards and samples as directed in the previous sections. Please predict the concentration before assaying. If values for these are not within the range of the standard curve, users must determine the optimal sample dilutions for their experiments. We recommend running all samples in duplicate.
| Step | |
| 1. | Add Sample: Add 100µL of Standard, Blank, or Sample per well. The blank well is added with Sample diluent. Solutions are added to the bottom of micro ELISA plate well, avoid inside wall touching and foaming as possible. Mix it gently. Cover the plate with sealer we provided. Incubate for 120 minutes at 37°C. |
| 2. | Remove the liquid from each well, don't wash. Add 100µL of Detection Reagent A working solution to each well. Cover with the Plate sealer. Gently tap the plate to ensure thorough mixing. Incubate for 1 hour at 37°C. Note: if Detection Reagent A appears cloudy warm to room temperature until solution is uniform. |
| 3. | Aspirate each well and wash, repeating the process three times. Wash by filling each well with Wash Buffer (approximately 400µL) (a squirt bottle, multi-channel pipette,manifold dispenser or automated washer are needed). Complete removal of liquid at each step is essential. After the last wash, completely remove remaining Wash Buffer by aspirating or decanting. Invert the plate and pat it against thick clean absorbent paper. |
| 4. | Add 100µL of Detection Reagent B working solution to each well. Cover with the Plate sealer. Incubate for 60 minutes at 37°C. |
| 5. | Repeat the wash process for five times as conducted in step 3. |
| 6. | Add 90µL of Substrate Solution to each well. Cover with a new Plate sealer and incubate for 10-20 minutes at 37°C. Protect the plate from light. The reaction time can be shortened or extended according to the actual color change, but this should not exceed more than 30 minutes. When apparent gradient appears in standard wells, user should terminatethe reaction. |
| 7. | Add 50µL of Stop Solution to each well. If color change does not appear uniform, gently tap the plate to ensure thorough mixing. |
| 8. | Determine the optical density (OD value) of each well at once, using a micro-plate reader set to 450 nm. User should open the micro-plate reader in advance, preheat the instrument, and set the testing parameters. |
| 9. | After experiment, store all reagents according to the specified storage temperature respectively until their expiry. |
When carrying out an ELISA assay it is important to prepare your samples in order to achieve the best possible results. Below we have a list of procedures for the preparation of samples for different sample types.
| Sample Type | Protocol |
| Serum | If using serum separator tubes, allow samples to clot for 30 minutes at room temperature. Centrifuge for 10 minutes at 1,000x g. Collect the serum fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. If serum separator tubes are not being used, allow samples to clot overnight at 2-8°C. Centrifuge for 10 minutes at 1,000x g. Remove serum and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. |
| Plasma | Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples at 4°C for 15 mins at 1000 × g within 30 mins of collection. Collect the plasma fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. Note: Over haemolysed samples are not suitable for use with this kit. |
| Urine & Cerebrospinal Fluid | Collect the urine (mid-stream) in a sterile container, centrifuge for 20 mins at 2000-3000 rpm. Remove supernatant and assay immediately. If any precipitation is detected, repeat the centrifugation step. A similar protocol can be used for cerebrospinal fluid. |
| Cell culture supernatant | Collect the cell culture media by pipette, followed by centrifugation at 4°C for 20 mins at 1500 rpm. Collect the clear supernatant and assay immediately. |
| Cell lysates | Solubilize cells in lysis buffer and allow to sit on ice for 30 minutes. Centrifuge tubes at 14,000 x g for 5 minutes to remove insoluble material. Aliquot the supernatant into a new tube and discard the remaining whole cell extract. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C. |
| Tissue homogenates | The preparation of tissue homogenates will vary depending upon tissue type. Rinse tissue with 1X PBS to remove excess blood & homogenize in 20ml of 1X PBS (including protease inhibitors) and store overnight at ≤ -20°C. Two freeze-thaw cycles are required to break the cell membranes. To further disrupt the cell membranes you can sonicate the samples. Centrifuge homogenates for 5 mins at 5000xg. Remove the supernatant and assay immediately or aliquot and store at -20°C or -80°C. |
| Tissue lysates | Rinse tissue with PBS, cut into 1-2 mm pieces, and homogenize with a tissue homogenizer in PBS. Add an equal volume of RIPA buffer containing protease inhibitors and lyse tissues at room temperature for 30 minutes with gentle agitation. Centrifuge to remove debris. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C. |
| Breast Milk | Collect milk samples and centrifuge at 10,000 x g for 60 min at 4°C. Aliquot the supernatant and assay. For long term use, store samples at -80°C. Minimize freeze/thaw cycles. |
