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  2. Life Science
  3. Metabolism Research
  4. ELISA / Assay Kits (Metabolism)
  5. Cholesterol Assay Kits
  6. LabAssay (TM) Cholesterol

LabAssay (TM) Cholesterol

for Cellbiology
Manufacturer :
FUJIFILM Wako Pure Chemical Corporation
Storage Condition :
Keep at 2-10 degrees C.
GHS :
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SDS
Comparison
Product Number
Package Size
Price
Inventory
Distributor
293-93601
Barcode No
4548995103437
500Tests
List Price
JPY 22,500

In stock in Japan

Please check here for notes on products and prices.

Document

SDS
Product Specification Sheet
Package Insert
Spectral Data
Certificate of Analysis
Calibration Certificate
Analytical Charts

Kit component

For 500 tests

Buffer Solution 150 mL x 1 bottle
Chromogen Substrate for 150 mL x 1 bottle
Standard Solution 5 mL x 1 bottle

Product Overview

Cholesterol concentration in serum is known to be closely related with production, absorption and dissimilation in liver and intestinal tract, and lipoprotein metabolism in blood.
LabAssay™ Cholesterol is a kit designed to measure total cholesterol in samples by an enzyme reaction using N-Ethyl-N-(2-hydroxy-3-sulfopropyl)-3,5-dimethoxyaniline sodium salt (DAOS).

[Note] LabAssay™ series are reagents for research purposes. They cannot be used for diagnostic purposes.

Kit Performance

Analysis sample Human Serum/Plasma
Mouse Serum/Plasma
Rat Serum/Plasma
Dog Serum/Plasma
Cat Serum/Plasma
Culture medium (D-MEM)*
Calibration curve range 50 - 592.2 mg/dL
Sample volume 2 μL
Measurement duration Approx. 10 min
Wavelength Primary wavelength 600 nm
Reference wavelength 700 nm
*It is tested by using the samples that standard substances have been added to the culture medium.
Measurement availability depends on the culture medium, cell type and culture conditions. Please perform a spiked recovery test or dilution linearity test in advance using the culture medium to be used in your experiment.

Example of Calibration Curve

Example of Calibration Curve

Assay principle

Cholesterol esters in the sample are decomposed into free cholesterol and fatty acid by cholesterol esterase, when the sample reacts with chromogen reagent. The cholesterol is oxidized with existing free cholesterol by cholesterol oxidase, and simultaneously hydrogen peroxide is produced. The produced hydrogen peroxide let DAOS and 4-Aminoantipyrin oxidize and condensate quantitatively by peroxidase (HRP), which produces a blue pigment. Quantitation of total cholesterol in the sample can be made by measurement of the absorbance.

Procedure

Assay in a microplate
Perform the assay in the wells according to the following table scheme.

Test Standard Blank
Chromogen reagent 300 μL 300 μL 300 μL
Sample 2 μL 2 μL -
Mix well and incubate at 37 ℃ for 5 min. Measure the absorbance of the test sample and standard solution with the blank solution as the control. Primary wavelength 600 nm / Referrence wavelength 700 nm.

Data

Repeatability (within run precision)

Repeatability was conducted by measuring sixfold assays of serum samples from human, mouse and rat at two concentrations with this product.

n＼ID Human serum Mouse serum Rat serum
ID1 (mg/dL) ID2 (mg/dL) ID1 (mg/dL) ID2 (mg/dL) ID1 (mg/dL) ID2 (mg/dL)
1 169.8 81.3 146.3 97.3 134.9 78.9
2 172.8 86.6 143.1 95.9 135.0 75.3
3 176.3 90.5 139.5 92.4 140.1 74.6
4 183.8 86.8 138.8 85.8 132.3 70.0
5 178.2 84.1 137.4 90.3 130.9 76.7
6 174.6 88.1 139.8 86.6 133.7 77.7
mean 175.9 86.2 140.8 91.4 134.5 75.5
SD 4.824 3.19 3.280 4.73 3.169 3.13
CV(%) 2.7 3.7 2.3 5.2 2.4 4.1

[Result]
The CV (%) of human serum was 2.7-3.7%, mouse serum was 2.3-5.2% and rat serum was 2.4-4.1%, indicating good repeatability.

Reproducibility (between run precision)

Reproducibility was conducted by measuring serum samples from human, mouse and rat at three concentrations each for four days with this product.

Day＼ID Human serum Mouse serum Rat serum
ID3 (mg/dL) ID4 (mg/dL) ID5 (mg/dL) ID3 (mg/dL) ID4 (mg/dL) ID5 (mg/dL) ID3 (mg/dL) ID4 (mg/dL) ID5 (mg/dL)
1 371.1 173.8 79.8 287.6 139.7 89.1 272.4 135.5 71.0
2 390.5 181.0 81.4 298.3 141.4 95.7 284.5 145.7 77.0
3 380.4 176.9 79.0 284.5 139.4 88.6 274.2 136.3 74.7
4 385.8 180.2 85.2 293.6 142.0 90.1 277.0 135.9 79.5
mean 382.0 178.0 81.4 291.0 140.6 90.9 277.0 138.4 75.6
SD 7.212 2.859 2.38 5.335 1.101 2.84 4.616 4.253 3.13
CV(%) 1.9 1.6 2.9 1.8 0.8 3.1 1.7 3.1 4.1

[Result]
The CV (%) of human serum was 1.6-2.9%, mouse serum was 0.8-3.1% and rat serum was 1.7-4.1%, indicating good reproducibility.

Dilution Linearity Test

Serum and plasma (EDTA) from human, mouse and rat, as well as D-MEM containing 10% FBS spiked with the standard, were serially diluted twofold with saline and measured in duplicate to assess linearity.

human serum
mouse serum
rat serum
D-MEM/10% FBS medium

[Result]
All measurement results showed good linearity within the measurement range.

Spike and Recovery Test

Standard solutions at four concentrations were added to serum/plasma (EDTA) from human, mouse and rat as well as D-MEM containing 10% FBS. Next, spike recovery tests were conducted, with each measurement performed in duplicate.

Human sample

Spiked amount
(mg/dL)		 Measurement value
(mg/dL)		 Recovery volume
(mg/dL)		 Recovery rate
(%)
Serum - 166.4 - -
47.6 219.1 52.6 111
99.7 276.1 109.7 110
150.4 326.3 159.9 106
201.3 378.3 211.9 105
Average 108
Plasma
(EDTA)		 - 130.2 - -
47.6 181.0 50.8 107
99.7 237.8 107.6 108
150.4 280.5 150.3 100
201.3 343.2 213.0 106
Average 105

Mouse sample

Spiked amount
(mg/dL)		 Measurement value
(mg/dL)		 Recovery volume
(mg/dL)		 Recovery rate
(%)
Serum - 86.1 - -
47.6 136.1 50.0 105
99.7 188.6 102.5 103
150.4 239.3 153.2 102
201.3 288.1 202.0 100
Average 103
Plasma
(EDTA)		 - 35.5 - -
47.6 84.5 49.1 103
99.7 134.4 98.9 99.2
150.4 188.9 153.5 102
201.3 230.6 195.1 96.9
Average 100

Rat sample

Spiked amount
(mg/dL)		 Measurement value
(mg/dL)		 Recovery volume
(mg/dL)		 Recovery rate
(%)
Serum - 69.9 - -
47.6 121.5 51.6 108
99.7 161.8 91.9 92.2
150.4 220.8 150.8 100
201.3 276.8 206.9 103
Average 101
Plasma
(EDTA)		 - 42.6 - -
47.6 91.8 49.2 106
99.7 137.6 95.0 105
150.4 180.1 137.5 98.3
201.3 235.8 193.2 98.6
Average 102

D-MEM

Spiked amount
(mg/dL)		 Measurement value
(mg/dL)		 Recovery volume
(mg/dL)		 Recovery rate
(%)
50% culture medium
＋standard solution		 - 3.22 - -
80.0 84.3 81.1 101
122.5 127.9 124.7 102
157.1 163.3 160.1 102
197.1 199.1 195.9 99.4
Average 101

[Result]
Good recovery rates were confirmed.

Measurement Using Actual Samples

Human sample

ID＼n Measurement value (mg/dL) mean SD CV
(%)
1 2
Serum ID1 191.7 173.6 182.7 12.80 7.0
Plasma ID1 143.2 143.1 143.2 0.07071 0.0

Rat sample (S.D. rat)

ID＼n Measurement value (mg/dL) mean SD CV
(%)
1 2
Serum ID1 60.0 67.1 63.6 5.02 7.9
ID2 84.6 89.0 86.8 3.11 3.6
ID3 90.7 80.8 85.8 7.00 8.2
ID4 82.1 72.3 77.2 6.93 9.0
ID5 89.3 93.9 91.6 3.25 3.6
Plasma
(EDTA)		 ID1 42.9 38.6 40.8 3.04 7.5
ID2 59.3 54.3 56.8 3.54 6.2
ID3 51.1 46.3 48.7 3.39 7.0
ID4 45.2 40.2 42.7 3.54 8.3
ID5 59.2 63.7 61.5 3.18 5.2

Mouse sample (CD-1(ICR))

ID＼n Measurement value (mg/dL) mean SD CV
(%)
1 2
Serum ID1 95.4 89.3 92.4 4.31 4.7
ID2 85.3 84.5 84.9 0.566 0.7
ID3 68.6 71.0 69.8 1.70 2.4
ID4 80.9 73.6 77.3 5.16 6.7
ID5 64.8 66.8 65.8 1.41 2.1
Plasma
(EDTA)		 ID1 41.2 39.1 40.2 1.48 3.7
ID2 43.7 40.8 42.3 2.05 4.9
ID3 35.8 38.7 37.3 2.05 5.5
ID4 43.1 41.4 42.3 1.20 2.8
ID5 39.3 40.7 40.0 0.990 2.5

Dog sample (Beagle)

ID＼n Measurement value (mg/dL) mean SD CV
(%)
1 2
Serum ID1 155.7 144.1 149.9 8.202 5.5
ID2 95.2 95.2 95.2 0.00 0.0
ID3 127.4 119.2 123.3 5.798 4.7
ID4 112.9 120.4 116.7 5.303 4.5
ID5 225.1 240.8 233.0 11.10 4.8
ID6 119.7 115.1 117.4 3.253 2.8
ID7 110.9 97.6 104.3 9.405 9.0
ID8 102.5 100.5 101.5 1.414 1.4
Plasma
(EDTA)		 ID1 146.6 135.8 141.2 7.637 5.4
ID2 119.7 130.2 125.0 7.425 5.9
ID3 157.1 164.6 160.9 5.303 3.3
ID4 122.2 133.9 128.1 8.273 6.5
ID5 132.2 129.4 130.8 1.980 1.5
ID6 133.8 154.3 144.1 14.50 10.1
ID7 117.4 119.5 118.5 1.485 1.3
ID8 103.6 111.6 107.6 5.657 5.3

Cat sample (Short hair)

ID＼n Measurement value (mg/dL) mean SD CV
(%)
1 2
Serum ID1 108.1 110.5 109.3 1.697 1.6
ID2 80.4 81.9 81.2 1.06 1.3
ID3 97.5 96.5 97.0 0.707 0.7
ID4 105.1 107.3 106.2 1.556 1.5
ID5 98.0 102.0 100.0 2.828 2.8
ID6 103.0 99.8 101.4 2.263 2.2
ID7 129.5 128.3 128.9 0.8485 0.7
ID8 71.0 70.4 70.7 0.424 0.6
Plasma
(EDTA)		 ID1 88.5 86.6 87.6 1.34 1.5
ID2 53.8 54.7 54.3 0.636 1.2
ID3 78.0 77.9 78.0 0.0707 0.1
ID4 73.3 75.1 74.2 1.27 1.7
ID5 80.4 80.9 80.7 0.354 0.4
ID6 60.0 60.4 60.2 0.283 0.5
ID7 71.5 72.4 72.0 0.636 0.9
ID8 87.5 90.6 89.1 2.19 2.5

FAQ

About calibration curve

Are there any precautions to consider when generating the calibration curve?
When purified water is measured as a sample, the absorbance should be 0.11 or less. When a 200 mg/dL cholesterol standard solution is measured, the absorbance should range from 0.11 to 0.65. If the absorbance values differ significantly from these criteria, repeat the measurement.

About sample

Is there any influence from substances that interfere with measurement (ascorbic acid, bilirubin, etc.)?
Ascorbic acid and bilirubin have almost no effect on the measured values.

About kit usage

What instruments, and equipment are required for the assay using this kit?
The instruments and equipment required for the use of this kit are listed below.
  • 96 well microplate (transparent type)
  • Micropipette
  • Plate mixer*
  • Incubator maintaining at 37°C*
  • Microplate reader with 600 nm wavelength filter

(*This is not necessary depending on the model of the microplate reader.)

(For Test Tube method)

  • Test tube
  • Pipette
  • Incubator maintaining at 37°C
  • Spectrophotometer or colorimeter with 600 nm wavelength filter
How should I use the standard solution?
Use the supplied standard solution as is or dilute it as necessary to prepare five standard concentrations according to the instructions provided in the “Package Insert” available in the Document section of this page.
What are the storage conditions and shelf-life guidelines for each reagent?
Chromogen Reagent :
Chromogen reagent is prepared by dissolving 1 vial of Chromogen Substrate with 1 vial of Buffer Solution. After adjustment, it may be stored at 2°C - 10°C for up to 3 weeks.

Overview / Applications

Property

Manufacturer Information

Alias

  • LabAssay
    635-50981 (Previous code)
    LABCHO-M1 (Previous code)
    294-65801 (Previous code)

For research use or further manufacturing use only. Not for use in diagnostic procedures.

Product content may differ from the actual image due to minor specification changes etc.

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