Name: Dog MAU (Microalbuminuria) ELISA Kit (AEKE12436)
Brand: Assay Genie
SKU: AEKE12436
Price: 649 EUR
Availability: InStock

Product Name:	Dog MAU (Microalbuminuria) ELISA Kit
SKU:	AEKE12436
Size:	96 Assays
Reactivity:	Canine
Assay Type:	Sandwich
Sensitivity:	1.45 µg/mL
Range:	3.13-200 µg/mL
Standard:	200 µg/mL
Assay Length:	3.5h
Sample Type:	Serum, plasma, tissue homogenates, cell lysates, cell culture supernates and other biological fluids.

Kit Components:

Component	Quantity		Storage
	48T	96T
Pre-Coated Microplate	6 strips x 8 wells	12 strips x 8 wells	4°C/-20°C
Standard (Lyophilized)	1 vial	2 vials	4°C/-20°C
Biotinylated Antibody (100x)	60 μL	120 μL	4°C/-20°C
Streptavidin-HRP (100x)	60 μL	120 μL	4°C/-20°C
Standard/Sample Diluent Buffer	10 mL	20 mL	4°C/-20°C
Biotinylated Antibody Diluent	6 mL	12 mL	4°C/-20°C
HRP Diluent	6 mL	12 mL	4°C/-20°C
Wash Buffer (25x)	10 mL	20 mL	4°C/-20°C
TMB Substrate Solution	6 mL	10 mL	4°C/-20°C (store in dark)
Stop Reagent	3 mL	6 mL	4°C/-20°C
Plate Covers	1 piece	2 pieces	-
Bradford Reagent	1 vial (2 mL)		RT

The Assay Genie Dog MAU (Microalbuminuria) ELISA Kit is a highly sensitive assay for the quantitative measurement of Dog MAU in the following samples: serum, plasma and other biological samples. This kit utilizes a sandwich enzyme-linked immunosorbent assay (ELISA) format. An antibody specific to the target protein is pre-coated onto the wells of a 96-well microplate. Samples and standards are added to the wells, allowing the target to bind to the immobilized antibody. After incubation, unbound substances are removed through washing. A biotinylated detection antibody is then added, which binds specifically to the captured target. Following a second wash to remove excess detection antibody, HRP-conjugated Streptavidin is introduced, forming a biotin-streptavidin-HRP complex. After a third washing step, TMB substrate is added to initiate a colorimetric reaction catalysed by HRP. The reaction produces a blue product that turns yellow upon addition of the acidic Stop Solution. The optical density (OD) is measured at 450 nm using a microplate reader. The OD450 value is directly proportional to the concentration of the target analyte in the sample, which can be determined by referencing a standard curve.

This dual function kit includes validated Bradford Reagent to quantify total protein concentration for accurate sample normalization.

Specificity:

This assay has high sensitivity and excellent specificity for detection of Dog MAU. No significant cross-reactivity or interference between Dog MAU and other targets was observed.

Linearity:

Matrix	1:2	1:4	1:8	1:16
Serum (n=5)	85-96%	82-88%	85-93%	78-89%
EDTA Plasma (n=5)	78-89%	91-104%	94-106%	94-106%
Heparin Plasma (n=5)	96-102%	87-98%	96-108%	91-104%

Recovery:

Matrix	Recovery Range	Average
Serum (n=5)	86-97%	92%
EDTA Plasma (n=5)	79-92%	86%
Heparin Plasma (n=5)	80-86%	83%

Precision:

Intra-assay Precision (Precision within an assay)：CV%<8%
Three samples of known concentration were tested twenty times on one plate to assess intra-assay precision.
Inter-assay Precision (Precision between assays)：CV%<10%
Three samples of known concentration were tested in forty separate assays to assess inter-assay precision.

*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.

Step	Protocol
1.	Reagent Preparation: Before adding to the wells, equilibrate the TMB substrate for at least 30 mins at room temperature. When diluting samples and reagents, ensure they are mixed completely and evenly. It is recommended to plot a standard curve for each test.
2.	Plate Setup: Set standard, test sample and control/blank (zero) wells on the pre-coated plate and record their positions. It is recommended to measure each standard and sample in duplicate.
3.	Standard, Samples & Control Loading: Aliquot 100 µl of standard working solution or samples and controls into the designated wells. Add all solutions carefully to the bottom of each micro-ELISA plate well. Avoid touching the inner walls of the wells and minimize foaming during pipetting.
4.	First Incubation: Seal the plate with a cover and incubate at 37°C for 80 mins.
5.	Washing: Remove the cover, aspirate the liquid from the plate. Wash plate with 200 µl Wash Solution and let it sit for 1-2 minutes. Remove the remaining liquid from all wells completely by snapping the plate onto absorbent paper. Wash plate 3 times. After the last wash, remove any remaining Wash Solution by aspirating or decanting. Invert the plate and blot it against absorbent paper.
6.	Biotinylated Antibody Working Solution Addition: Add 100 µl of Biotin-labelled Antibody working solution to the bottom of each well (standard, test sample & zero wells) without touching the side walls.
7.	Second Incubation: Seal the plate with a cover and incubate at 37°C for 50 mins.
8.	Washing: Remove seal and aspirate. Repeat wash process for total 3 times as conducted in step 5.
9.	Addition Streptavidin-HRP Working Solution: Add 100 µl of Streptavidin-HRP Working Solution into each well, cover the plate and incubate at 37°C for 50 mins.
10.	Washing: Remove seal and aspirate. Repeat wash process for total 5 times as conducted in step 5.
11.	TMB Substrate Addition and Colour Development: Add 90 µl of TMB substrate into each well, cover the plate and incubate at 37°C in dark for 20 mins. (Note: Do NOT exceed 30 minutes.) As soon as a blue colour develops preheat the Microplate Reader.
12.	Stop Reaction: Add 50 µl of Stop solution into each well and mix thoroughly. The colour changes into yellow immediately. Note: If colour change does not appear uniform, gently tap the plate to ensure thorough mixing.
13.	OD Measurement: Read the O.D. absorbance at 450 nm in a microplate reader immediately after adding the stop solution.

The procedures outlined below are provided as general recommendations for sample preparation in ELISA assays. Due to the variability of biological samples and specific assay requirements, users are advised to optimize protocols based on their own experimental conditions.

Sample Type	Protocol
Serum	Allow whole blood to coagulate at room temperature (2 h) or 2-8°C overnight. Centrifuge at 1000 × g for 20 min and collect the supernatant. Store or use immediately.
Plasma	Collect in anticoagulant tubes (EDTA, citrate, or heparin), mix gently, and centrifuge within 30 min at 1000 × g, 2-8°C for 15 min. Store or assay as needed.
Tissue homogenates	Avoid buffers containing NP-40, Triton X-100, or DTT, as these strongly inhibit the assay. We recommend using 50 mM Tris + 0.9% NaCl + 0.1% SDS, pH 7.3. 1. Place the target tissue on ice, rinse with pre-cooled PBS buffer (0.01 M, pH 7.4) to remove residual blood and weigh the tissue. 2. Homogenize the tissue on ice using an appropriate lysis buffer. The lysate volume should correspond to the tissue weight; typically, 9 mL PBS is used per 1 g of tissue. It is recommended to add protease inhibitors to the PBS (e.g., 1 mM PMSF). 3. Further disrupt the tissue using ultrasonic homogenization (keep samples on an ice bath during sonication) or freeze-thaw cycles (repeat twice for effective lysis). 4. Centrifuge the homogenate at 5000 × g for 5 minutes. Collect the supernatant for immediate analysis, or aliquot and store at -20°C or -80°C for future assays. 5. Determine total protein concentration using the Bradford Reagent included in this kit. For ELISA assays, the total protein concentration should generally be 1-3 mg/mL. Note: Tissues with high endogenous peroxidase levels (e.g., liver, kidney, pancreas) may react with TMB substrate, causing false positives. If this occurs, treat samples with 1% H₂O₂ for 15 minutes before repeating the assay.
Cell culture supernatant	Centrifuge the sample at 2500 rpm for 5 minutes at 2-8°C. Carefully collect the clarified cell culture supernatant for immediate analysis, or aliquot and store it at -80°C for future assays.
Suspension cell lysates	Centrifuge the cell suspension at 2500 rpm for 5 minutes at 2-8°C and collect the cell pellet. Wash the pellet with pre-cooled PBS (0.01 M, pH 7.4) and mix gently. Repeat centrifugation and discard the supernatant. Add 0.5-1 mL of cell lysis buffer containing an appropriate protease inhibitor (e.g., PMSF, final concentration: 1 mM). Lyse the cells on ice for 30-60 minutes or disrupt them using ultrasonic homogenization. Centrifuge the lysate at 10,000 rpm for 10 minutes at 2-8°C and collect the supernatant for immediate use, or aliquot and store at -80°C.
Adherent cell lysates	Remove the supernatant and wash the cells three times with pre-cooled PBS. Add 0.5-1 mL of cell lysis buffer supplemented with an appropriate protease inhibitor (e.g., PMSF at a final concentration of 1 mmol/L). Scrape the adherent cells using a cell scraper and transfer the cell suspension to a centrifuge tube. Lyse the cells on ice for 30-60 minutes, or disrupt the cells by ultrasonic treatment. Centrifuge the lysate at 10,000 rpm for 10 minutes at 2-8°C and collect the supernatant for immediate use, or aliquot and store at -80°C.
Sample storage	Short-term: 2-8°C for up to 5 days. Medium-term: -20°C for up to 6 months. Long-term: -80°C or cryopreservation in liquid nitrogen. Frozen samples should be thawed rapidly in a 15-25°C water bath to minimize ice crystal-induced damage. Thawed samples can be analyzed immediately or stored temporarily at 2-8°C. Repeated freeze-thaw cycles should be strictly avoided due to their detrimental effect on protein stability.
Other sample types	For more information about how to process other sample types (e.g., body fluids, breast milk & more), please contact our Tech Support Team at techsupport@assaygenie.com.