Purple-Jelley Hyaluronic Acid assay kit

Biocolor’s Purple-Jelley assay kit is the perfect tool for accurate measurement of hyaluronic acid / Hyaluronan levels in your samples. This colorimetric assay is optimised for quantitative analysis in-vivo, tissue-derived hyaluronic acid / Hyaluronan and includes full step-by-step instructions.

Colorimetric Detection (655nm) (Endpoint)

Hyaluronic Acid: A gentle giant!

Hyaluronic acid, in its hydrated form, is a unique carbohydrate polymer, often referred to as a ‘gentle giant.’ It consists of a lengthy, flexible, non-branching chain with a repeating disaccharide pattern. This disaccharide is composed of alternating uronic acid and aminosugar units.

Why is our kit called ‘Purple-Jelley’?

Discovering the J-Aggregate Effect in Cyanine DyesIn 1936, Edwin Jelley made a fascinating observation, documented it in a letter to Nature (Nature 138, 1009 – 1010). He noted a peculiar behaviour of certain cyanine dyes, that when dissolved in 5 M NaCl, they dyes exhibited a third absorbance peak at a longer wavelength, around 650nm. In deionized water, however, they displayed only a double peak at approximately 540nm and 570nm. The 650nm peak in concentrated dye solutions resulted from the aggregation of dye molecules and was later termed a ‘J-aggregate,’ in honor of Edwin Jelley. The J-aggregate is known as a supra-molecular complex, formed by stacking individual dye molecules.

Subsequent research in the 1960s, notably by Kay et al. (J. Physical Chem. 68, 1896 – 1906), revealed that various biological polymers, including proteins, DNA, polar lipids, and glycosaminoglycans, could also induce this third absorbance peak. This phenomenon led to the development of the Purple-Jelley assay, named after the purple color of the dye reagent and Edwin Jelley himself.

‍

An overview of the Purple-Jelley assay steps:

During the assay, hyaluronic acid is selectively purified during the assay sample preparation protocol. This is then reacted with the Purple-Jelley dye reagent, and the absorption of the characteristic third wavelength recorded. By comparison with a calibration curve the hyaluronic acid content of the sample can be measured.

‍Step 1. The assay protocol takes tissue samples through a sequential sample preparation protocol which involves enzymatic protein digestion, followed by precipitation and purification of GAGs, culminating in the precipitation of purified Hyaluronic acid.

Step2. The processed sample is then incubated for 10 minutes with the Purple-Jelley dye reagent, forming a coloured product which can be measured spectrophotometrically.

Step 3. The Hyaluronic acid content of unknown samples can be calculated by comparison against a calibration curve prepared using a standard comprising hyaluronic acid (supplied with the kit).

‍

Assay range

10 – 100µg/ml

Limit of Detection

10µg/ml

Detection Method

Colorimetric Detection (655nm) (Endpoint)

Measurements per kit

100 in total (allows a maximum of 46 samples to be run in duplicate alongside a standard curve).

Suitable Samples

In-vivo: Hyaluronic acid purified from in-vivo tissues. The kit protocol involves extraction and purification of hyaluronic acid prior to reaction with the Purple-Dye reagent.

Precautions

This kit is designed for research use only. Not for use in diagnostic procedures.
Kit requires access to a centrifuge, as well as a spectrophotometer/colorimeter capable of colorimetric, absorbance detection at 655nm.
Specific sample preparation protocols may require customer to provide further reagents, consult assay manual for further information.

ation

Mode of ActionAssay SpecificationsKit Contents

Purple-Jelley Hyaluronic acid kit contents:‍

1. Purple-Jelley Dye Reagent (1x 20ml)

2. Hyaluronan Reference Standard (1x 5ml, 0.2mg/ml soluble Hyaluronic Acid)

3. Precipitating Reagent (2x 34ml)

4. Sodium Chloride (1x 20ml)

5. Cetylpyridinium Chloride (1x 20ml)

6. TRIS-buffered Saline (5x tablets)

7. 2ml screw-cap tubes for preparation of samples.

8. Assay kit manual

NB: Additional reagents may be required for sample preparation prior to assay. Consult manual or contact us for further details.

Biocolor’s Purple-Jelley assay kit is the perfect tool for accurate measurement of hyaluronic acid / Hyaluronan levels in your samples. This colorimetric assay is optimised for quantitative analysis in-vivo, tissue-derived hyaluronic acid / Hyaluronan and includes full step-by-step instructions.

Introduction

Blood samples contain rich DNA, including mitochondrial DNA, genomic DNA, circulating DNA (mostly released into blood after tumor cell apoptosis) in white blood cells, as well as parasitic viral or microbial DNA. These DNA are important parameters in clinical testing or diagnosis, which are also valuable materials for medical research. There are three main issues with extracting DNA from blood samples:

1. The sample is highly infectious, posing great harm to operators and the environment.

2. The source of DNA is complex and aportion of the nucleic acid, such as viral DNA or free DNA, may be lost during the operation, leading to downstream detection failure;

3. Blood sample contains a large amount of impurities and inhibitory factors.

Currently there are many methods available for extracting DNA from whole blood samples, such as phenol chloroform extraction, salting out method, etc. However, these methods require pre-treatment of blood sample, which removes red blood cells and isolate white blood cells in the first step. Due to the requirement that it cannot inactivate or kill pathogens during the process of removing red blood cells, the waste liquid (red blood cell lysate) and consumables may be contaminated by pathogens and become infectious, posing a danger to the entire laboratory environment and operators. In addition, during the process of removing red blood cells, useful nucleic acid information such as viruses, microorganisms, or circulating DNA is also lost, leading to experiment or detection failures.

The HiPure Blood DNA Kits series provided by Magen Company uses silica gel column purification technology, which can directly lyse whole blood samples without the need for white blood cell separation. Whole blood samples are directly mixed with lysates and proteases, resulting in the inactivation of pathogens, greatly reducing the infectivity, environmental pollution, and the chance of operators being infected. Due to the direct lysis and digestion of samples, except lymphocyte DNA, other circulating DNA as well as DNA from viruses and microorganisms, can also be recovered.

This product provides fast and easy methods for purification of total DNA for reliable PCR and Southern blotting. Total DNA (e.g., genomic, viral, mitochondrial) can be purified from whole blood, tissue and culture cells.

Details

Specifications

Features	Specifications
Main Functions	Isolation total DNA from 10ml blood and 1g tissue using Maxi column
Applications	PCR, southern bolt and virus detection, etc
Purification method	Maxi spin column
Purification technology	Silica technology
Process method	Manual (centrifugation or vacuum)
Sample type	Tissue, cell, blood, saliva, swab, blood spot, semen and other clinical samples
Sample amount	3-10ml
Elution volume	≥700μl
Time per run	≤90 minutes
Liquid carrying volume per column	4ml
Binding yield of column	5mg

Principle

This product is based on silica column purification. The sample is lysed and digested with lysate and protease, DNA is released into the lysate. Transfer to an adsorption column. Nucleic acid is adsorbed on the membrane, while protein is not adsorbed and is removed with filtration. After washing proteins and other impurities, Nucleic acid was finally eluted with low-salt buffer (10mm Tris, pH9.0, 0.5mm EDTA).

Advantages

• High quality DNA – meet a variety of downstream applications, including PCR, qPCR, enzyme digestion, hybridization, etc.
• Fast – without separation of leukocytes, organic extraction or ethanol precipitation
• Simple – all nucleic acids can be obtained by direct digestion
• Pertinence – specially designed for isolating DNA from 3-10ml blood and related body fluids
• Wide applicability – handle a variety of liquid samples
• Economy – excellent cost effectiveness performance

Kit Contents

Contents	D311502	D311503
Purification Times	10	50
HiPure gDNA Maxi Columns	10	50
50ml Collection Tubes	20	100
Buffer ATL	120 ml	550 ml
Buffer AL	120 ml	2 x 300 ml
Buffer GW1*	53 ml	220 ml
Buffer GW2*	25 ml	110 ml
RNase A	40 mg	180 ml
Proteinase K	120 mg	540 mg
Protease Dissolve Buffer	12 ml	50 ml
Buffer AE	30 ml	120 ml

Storage and Stability

Proteinase K, RNase A should be stored at 2-8°C upon arrival. However, short-term storage (up to 12 weeks) at room temperature (15-25°C) does not affect their performance. The remaining kit components can be stored at room temperature (15-25°C) and are stable for at least 18 months under these conditions.

Purchase Guide

Name	CAT NO	Sample amount	Leukocyte protocol*	Colum type	Elutio volume	Average yield	Time per run
HiPure Blood DNA Mini Kit	D3111	10-200μl	1ml	2ml column	≥20μl	5-9μg/200μl	≤30 minutes
HiPure Tissue&Blood DNA Midi Kit	D3113	0.2-2ml	10ml	1.5ml column	≥300μl	20-40μg/1m	≤80 minutes
HiPure Tissue&Blood DNA Maxi Kit	D3115	3 -10ml	10ml	15ml column	≥700μl	20-40μg/1m	≤90 minutes
HiPure Tissue&Blood DNA 96 Kit	D3117	1-200μl	1ml	96 well plate		3-8μg/200μl

Blood samples contain rich DNA, including mitochondrial DNA, genomic DNA, circulating DNA (mostly released into blood after tumor cell apoptosis) in white blood cells, as well as parasitic viral or microbial DNA. These DNA are important parameters in clinical testing or diagnosis, which are also valuable materials for medical research. There are three main issues with extracting DNA from blood samples:

Overview

• Removes primer dimers
• Simultaneously clean-up and normalize PCR products 
• Fast (less than 20 minutes), high-throughput and easy processing using centrifugation
• Sufficient elution volume (100 µL) for repeat or future assays 
• Non-magnetic bead purification 

Norgen’s NGS Normalization 96-Well Kit allows for high-throughput NGS library PCR amplicon clean-up and normalization of NGS library PCR product concentration. The clean-up removes all PCR by-products including primers, dimers, enzymes and unincorporated nucleotides during the library prep PCR step. Purification is based on 96-well column chromatography using Norgen’s proprietary resin as the separation matrix. The amount of Norgen’s matrix in each 96 well is optimized to have a limited binding capacity, thus each well can elute an equal concentration of PCR product. The process involves first adding 3 volumes of Buffer SK to the PCR product, and the mixture is then loaded onto the 96-Well Normalization Plate. The bound PCR amplicon is then washed with the provided Wash Solution A in order to remove any remaining impurities, and the purified PCR amplicon is eluted with Elution Buffer B. The purified and normalized library PCR amplicon can then be used in NGS workflows and other sequencing applications.

Details

Supporting Data

Figure 1 / 3

Previous

Next

Click for expanded view

Storage Conditions
All solutions should be kept tightly sealed and stored at room temperature. This kit is stable for 2 years after the date of shipment.

Component	Cat. 61900 (192 preps)
Buffer SK	30 mL
Wash Solution A	2 x 38 mL
Elution Buffer B	30 mL
96-Well Normalization Plate	2
96-Well Collection Plate	2
96-Well Elution Plate	2
Adhesive Tape	1
Product Insert	1