Human CDA/Cytidine deaminase ELISA Kit- High Sensitivity

Description

Description

Human CDA / Cytidine deaminase ELISA

CDA (Cytidine Deaminase) encoded protein is a cytosolic, zinc-containing enzyme that catalyzes the deamination of cytidine to uracil. This enzyme is necessary for the normal turnover of DNA and is required for the generation of new cells. Mutations in tCDA are associated with severe combined immunodeficiency (SCID), a condition characterized by a lack of both B and T cells. Diseases associated with CDA include Acute Leukemia and Immunodeficiency With Hyper-Igm, Type 2.

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:	Human CDA / Cytidine deaminase ELISA Kit
Product Code:	HUFI02139
Size:	96 Assays
Alias:	CDA, Cytidine aminohydrolase, CDD
Detection Method:	Sandwich ELISA, Double Antibody
Application:	This immunoassay kit allows for the in vitro quantitative determination of Human CDA concentrations in serum plasma and other biological fluids.
Sensitivity:	0.188ng/ml
Range:	0.313-20ng/ml
Storage:	4°C for 6 months
Note:	For Research Use Only

Additional Information

Recovery:

Matrices listed below were spiked with certain level of Human CDA and the recovery rates were calculated by comparing the measured value to the expected amount of Human CDA in samples.


Matrix	Recovery Range (%)	Average (%)
serum (n=5)	87-101	94
EDTA plasma (n=5)	91-105	97
UFH plasma (n=5)	87-105	95

Linearity:

The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Human CDA and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.


Sample	1:2	1:4	1:8
Serum (n=5)	87-100%	85-101%	92-102%
EDTA plasma (n=5)	83-100%	82-89%	83-99%
UFH Plasma (n=5)	81-96%	81-89%	81-94%

CV(%):

Intra-Assay: CV<8%
Inter-Assay: CV<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

Protein Information


Uniprot:	P32320
UniProt Protein Function:	CDA: This enzyme scavenge exogenous and endogenous cytidine and 2'-deoxycytidine for UMP synthesis. Belongs to the cytidine and deoxycytidylate deaminase family.
UniProt Code:	P32320
NCBI GenInfo Identifier:	1705718
NCBI Gene ID:	978
NCBI Accession:	P32320.2
UniProt Related Accession:	P32320

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 Type

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.

CDA Background

Cytidine deaminase (CDA) is an enzyme that plays a crucial role in nucleotide metabolism. It catalyzes the deamination of cytidine and deoxycytidine, converting them into uridine and deoxyuridine, respectively. This enzymatic reaction involves the removal of an amino group (-NH2) from the cytosine base of the nucleosides.

CDA's Role in Nucleotide Metabolism

CDA is found in various tissues and cell types, including the liver, kidney, gastrointestinal tract, lymphoid tissues, and bone marrow. It is particularly important in the context of nucleoside-based chemotherapy because it can inactivate certain nucleoside analog drugs commonly used in cancer treatment.

CDA FAQs

What is the CDA ELISA Kit?

The Human CDA / Cytidine Deaminase ELISA is an enzyme-linked immunosorbent assay designed to measure the levels of CDA in human samples.

What are the advantages of using the CDA ELISA Kit?

The Human CDA / Cytidine Deaminase ELISA provides accurate quantification of CDA levels, is user-friendly and easy to use, and delivers rapid results, saving time in the laboratory.

Where can I find more information about the CDA ELISA Kit?

For more detailed information about the CDA ELISA Kit, including technical specifications, performance characteristics, and ordering details, please refer to the product brochure or contact our customer support team. We are here to assist you with any inquiries you may have.

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Human CDA / Cytidine deaminase ELISA Kit

Description