Monkey IFN alpha/Interferon Alpha ELISA Kit

Description

Description

Monkey IFN alpha / Interferon Alpha ELISA Kit

Interferon alpha (IFN-alpha) is a type of protein that plays a crucial role in the immune system's response against viral infections and other immune challenges. IFN-alpha is a versatile molecule that exhibits antiviral and immunomodulatory properties, making it a valuable target for research in the field of immunology. By using the Monkey IFN alpha ELISA kit, researchers can gain insights into the immune response of monkeys to viral infections or other immune challenges. The ELISA kit provides a reliable and quantitative method to assess IFN-alpha levels, enabling researchers to better understand the role of IFN-alpha in monkey immune responses and potentially develop new therapeutic strategies.

Datasheet MSDS

Key Features

	Save Time	Pre-coated 96 well plate
	Quick Start	Kit includes all necessary reagents
	Publication Ready	Reproducible and reliable results

Overview


Product Name:	Monkey IFN-alpha (Interferon Alpha) ELISA Kit
Product Code:	MKFI00028
Size:	96 Assays
Alias:	IFN-alpha, Interferon Alpha
Detection Method:	Sandwich ELISA, Double Antibody
Reactivity:	Monkey
Sensitivity:	9.375pg/ml
Range:	15.625-1000pg/ml
Storage:	4°C for 6 months
Note:	For Research Use Only

Additional Information


Recovery	Matrices listed below were spiked with certain level of Monkey IFN-alpha and the recovery rates were calculated by comparing the measured value to the expected amount of Monkey IFN-alpha in samples. Please contact us for more information.
Linearity:	The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Monkey IFN-alpha and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected. Please get in contact for more information.
CV(%)	Intra-Assay <8 Inter-Assay <10

Kit Components

Component	Quantity	Storage
ELISA Microplate (Dismountable)	8x12 strips	4°C for 6 months
Lyophilized Standard	2	4°C/ -20°C
Sample/Standard Dlution Buffer	20ml	4°C
Biotin-labeled Antibody (Concentrated)	120ul	4°C (Protection from light)
Antibody Dilution Buffer	10ml	4°C
HRP-Streptavidin Conjugate (SABC)	120ul	4°C (Protect from light)
SABC Dilution Buffer	10ml	4°C
TMB Substrate	10ml	4°C (Protection from light)
Stop Solution	10ml	4°C
Wash Buffer (25X)	30ml	4°C
Plate Sealer	5	-

Other materials 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

Protocol

*Note: Protocols are specific to each batch/lot. For the exact instructions please follow the protocol included in your kit.

Before adding to wells, equilibrate the SABC working solution and TMB substrate for at least 30 min at 37°C. When diluting samples and reagents, they must be mixed completely and evenly. It is recommended to plot a standard curve for each test.

Step	Procedure
1.	Set standard, test sample and control (zero) wells on the pre-coated plate respectively, and then, record their positions. It is recommended to measure each standard and sample in duplicate. Wash plate 2 times before adding standard, sample and control (zero) wells!
2.	Aliquot 0.1ml standard solutions into the standard wells.
3.	Add 0.1 ml of Sample / Standard dilution buffer into the control (zero) well.
4.	Add 0.1 ml of properly diluted sample ( Human serum, plasma, tissue homogenates and other biological fluids.) into test sample wells.
5.	Seal the plate with a cover and incubate at 37 °C for 90 min.
6.	Remove the cover and discard the plate content, clap the plate on the absorbent filter papers or other absorbent material. Do NOT let the wells completely dry at any time. Wash plate X2.
7.	Add 0.1 ml of Biotin- detection antibody working solution into the above wells (standard, test sample & zero wells). Add the solution at the bottom of each well without touching the side wall.
8.	Seal the plate with a cover and incubate at 37°C for 60 min.
9.	Remove the cover, and wash plate 3 times with Wash buffer. Let wash buffer rest in wells for 1 min between each wash.
10.	Add 0.1 ml of SABC working solution into each well, cover the plate and incubate at 37°C for 30 min.
11.	Remove the cover and wash plate 5 times with Wash buffer, and each time let the wash buffer stay in the wells for 1-2 min.
12.	Add 90 µl of TMB substrate into each well, cover the plate and incubate at 37°C in dark within 10-20 min. (Note: This incubation time is for reference use only, the optimal time should be determined by end user.) And the shades of blue can be seen in the first 3-4 wells (with most concentrated standard solutions), the other wells show no obvious color.
13.	Add 50 µl of Stop solution into each well and mix thoroughly. The color changes into yellow immediately.
14.	Read the O.D. absorbance at 450 nm in a microplate reader immediately after adding the stop solution.

Sample Preparation

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.

Interferon Alpha Background

Interferon alpha / IFN-alpha background

Interferon alpha (IFN-alpha) is a type of protein that belongs to the family of interferons, which are signaling molecules released by cells in response to viral infections, certain cancers, and other immune triggers. IFN-alpha plays a vital role in the immune system's response against viral infections, as it helps to stimulate the immune cells, inhibit viral replication, and regulate various immune functions.

Interferon alpha gene

Interferon alpha (IFN-alpha) is encoded by a gene cluster located on chromosome 9 (location 9p21.3) in humans, known as the interferon alpha gene cluster. This cluster consists of multiple genes that produce different subtypes of IFN-alpha. These genes are regulated by various factors and are typically induced in response to viral infections or immune signaling. The subtypes of IFN-alpha exhibit slight variations in their amino acid sequences, contributing to differences in their biological activities and potencies.

The IFN-alpha gene cluster provides the genetic blueprint for the synthesis of various IFN-alpha subtypes. This allows for a diverse immune response against different pathogens and immune challenges. The expression of these genes is tightly regulated, and their products are important for activating the immune system's response to viral infections and other immune triggers.

Interferon alpha Receptor and signalling pathways involved

The Interferon Alpha (IFN-alpha) receptor is a protein complex that is involved in the signaling pathways triggered by IFN-alpha binding. It consists of two subunits: IFNAR1 (Interferon Alpha Receptor 1) and IFNAR2 (Interferon Alpha Receptor 2) and is ubiquitously produced.

When IFN-alpha is released in response to viral infection or immune stimulation, it binds to the IFN-alpha receptor on the surface of target cells (immune cells of virus-infected cells). Upon IFN-alpha binding, the receptor subunits undergo a conformational change, leading to the activation of intracellular signaling molecules. This activation typically involves the Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway.

The signaling pathways triggered by IFN-alpha receptor activation are diverse and can lead to various cellular responses. They include the induction of antiviral defense mechanisms, modulation of immune cell activity, and regulation of inflammatory responses.

In addition to the JAK-STAT pathway, IFN-alpha signaling can also involve other downstream signaling cascades, such as the mitogen-activated protein kinase (MAPK) pathway and the phosphoinositide 3-kinase (PI3K)/Akt pathway. These pathways contribute to the diverse biological effects of IFN-alpha on cell growth, differentiation, and immune responses.

Interferon alpha structure

IFN-alpha is a small protein composed of approximately 166 to 166 amino acids. It has a three-dimensional structure characterized by a bundle of alpha-helices. The protein forms dimers, meaning two identical IFN-alpha molecules come together to create a functional unit. The dimerization of IFN-alpha is essential for its biological activity.

Predicted structure of Interferon alpha in humans. Source:Uniprot

Interferon alpha (IFN alpha) Function

The primary function of IFN-alpha is to activate the immune response against viral infections. It acts by binding to specific receptors on the surface of target cells. Upon binding, it triggers a signaling cascade that activates various antiviral defense mechanisms within the cell. These mechanisms include the upregulation of genes involved in inhibiting viral replication, promoting the production of enzymes that degrade viral RNA, and enhancing the presentation of viral antigens to immune cells for recognition and elimination.

Interferon alpha (IFN alpha) In treatment

IFN-alpha is widely used in medicine for its antiviral and immunomodulatory properties. It has been employed in the treatment of chronic viral hepatitis B and C, certain types of leukemia, and various cancers, including melanoma and renal cell carcinoma. IFN-alpha is also used in the treatment of certain autoimmune disorders, such as multiple sclerosis and systemic lupus erythematosus, due to its ability to regulate immune responses.

However, the use of IFN-alpha is not without limitations and potential side effects. Its administration can lead to flu-like symptoms, including fever, fatigue, and muscle aches. Prolonged use of IFN-alpha may also have adverse effects on the bone marrow and the cardiovascular system. Nonetheless, IFN-alpha continues to be an important therapeutic agent in several medical conditions, and ongoing research aims to improve its efficacy and reduce side effects through modified formulations and targeted delivery methods.

Monkey Interferon Alpha FAQs

Q: What is the purpose of the Monkey IFN Alpha ELISA kit?

The Monkey Interferon Alpha ELISA kit is a laboratory test that allows researchers to measure the levels of Interferon Alpha (IFN-alpha) specifically in monkey samples.

Q: What type of samples can be used with the Monkey IFN Alpha ELISA kit?

The Monkey IFN Alpha ELISA kit is suitable for biological fluids such as serum samples

Q: Can the Monkey Interferon Alpha ELISA kit be used with other species?

No, the Monkey Interferon Alpha ELISA kit is specifically designed and validated for measuring IFN-alpha in monkey samples. It may not be suitable for other species due to differences in the amino acid sequences of IFN-alpha and potential variations in the performance of the kit. It is recommended to use species-specific ELISA kits for accurate measurements in different animal models.

Q: Where can I find additional technical support or assistance with the Monkey IFN Alpha ELISA kit?

For any technical inquiries or assistance regarding the Monkey IFN Alpha kit, you can reach out to our team. They will be available to answer your questions and provide the necessary guidance to ensure a successful experiment.

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