Calcium ( Ca ) Assay Kit LS ( CPZⅢ )
#CA02ME 250 T 35,800 JPY (excluding tax)
#CA01ME 500 T 43,000 JPY (excluding tax)
Measurement target
Testing sample
Measurement
range
Measurement wavelength (maximum wavelength)
Document
Ca2+
Serum, plasma,
urine, etc.
680 〜 700 nm (690 nm)
0.2 ～ 30 mg/dL
SDS
Instruction manual
・Calcium ( Ca ) Assay Kit LS ( CPZⅢ ) enables rapid quantification of calcium (Ca2+) in biological samples (serum, plasma, etc.) using a microplate reader (96 wells).
・The principle of this assay is a chelation quantification method, not an immunoassay, so it is species-independent.
・No pretreatment is required for serum or plasma samples.
・Multi-point calibration is not required as calibration is performed using two points along with the standard and blank sample included in the kit.
・ The kit does not contain any toxic or hazardous substances, so it is safe to handle and dispose of.
※ This product is for research use only. Do not use for diagnostic purposes.
Calcium accounts for about 1.5-2.9% of body weight and is an important element in the formation of the bones and soft tissues.
In the body, 90% of calcium is present in bones as hydroxyapatite, 1% in cells, and 0.1% in the blood. Physiological functions include osmotic pressure regulation in cells, blood coagulation, muscle atrophy, nerve stimulation, and enzyme activation. In blood, 45% of calcium is bound to albumin and globulin, 10% is phosphate, and 45% is free ions.
When serum calcium levels decrease, parathyroid hormone is secreted, which promotes bone resorption, reabsorption in the kidney, and calcium absorption from the small intestine associated with the production of active vitamin D, thereby increasing the serum calcium concentration. Through these mechanisms, serum calcium concentration is strictly regulated, and normal values remain within a narrow range.
Hypocalcemia is known to cause rickets, osteomalacia, tetany, osteoporosis, and arteriosclerosis, while hypercalcemia is known to be involved in depression, disorientation, fatigue, hypersensitivity, arrhythmia, and hypertension.
This method determines the calcium concentration by observing the color change in the visible region by the formation of a chelate complex between Chlorophosphonazo-Ⅲ and calcium.
Chlorophosphonazo-Ⅲ forms complexes with calcium in the neutral region. The calcium concentration can be determined by observing the absorbance at 690 nm as the main wavelength and 740-810 nm as the secondary wavelength.
◯ Samples that can be directly quantified with this kit
Examples: serum, plasma
The denaturant in the kit dissociates calcium from the protein and develops color after reducing agent reduces it. No specific pretreatment is required.
※ Do not use blood collection tubes containing EDTA which may affect the assay value.
◯ Samples that require dilution
Example: Urine
There is a possibility that the measurement range will be exceeded so dilute samples to 5x for measurement.
※ If using urine as a sample, preserve it by adding hydrochloric acid. When measuring, adjust the pH to 2~3.
※ The components of urine change from time to time, so the dilution ratio may vary. Be careful especially with the dilution ratio for early morning urine.
※ If you dilute a sample, be careful not to forget to multiply the dilution ratio when calculating the concentration.
◯ Samples that require acid extraction
Examples: cell lysates, tissue homogenates, and other aqueous solution samples
The pH of the samples was adjusted to low level to dissociates the calcium from the protein, and the supernatant was used as the assay sample. However, only the following chemicals can be measured.
・Free calcium
・Protein-bound (coordinated) calcium (Protein-Ca2+, e.g. albumin, metallothionein)
・Other coordination-bound calcium
※ Do not use lysis buffers containing EDTA as they may affect the measured values.
※ Acid digestion should be performed in the case of organic calcium.
◯ Samples that require microwave method and organic matter mixed acid decomposition
Example: Samples containing strong chelating agents such as EDTA, organic calcium (-C-Ca-C- single bonded), or calcium encapsulated in a cyclic ligand.
Acid extraction is necessary when measuring chemical species that cannot be dissociated or samples containing interfering substances such as strong calcium chelating agents.
※ After decomposition of organic matter, adjust the pH to 2-3, and use the sample as the assay sample.
(1) ) Add 2 µl of DIW, standard solution, and assay sample to wells of a 96-well plate.
(2) Add 240 µl of RA solution to each well
(3) Incubate the plate at room temperature for 10 minutes.
(4) Measure absorbance at main wavelength 690 nm (secondary wavelength 740-800 nm.)
(5) Calculate the calcium concentration using the following formula:
Intended use : | Measurement of calcium (Ca2+) |
Testing sample : | Serum, plasma, urine, cell lysates, tissue extract, plant extract, hair sample extract, food extract, drinking water, environmental water, mineral water, etc. |
Measurement range : | 0.2 ～30 mg/dL |
Measurement wavelength (maximum wavelength) : | 680 〜 700 nm（690 nm） |
Number of measurement: | 250 (CA02ME) / 500 (CA01ME) |
Assay method: | CPZIII method |
Species specificity: | Any species of organism is acceptable. |
Regulations (Poisonous and Deleterious Substances Control Act) | Not applicable |
Effects of coexisting substances: | The effect of 10 mg/dL of magnesium on the measurement value is within 5%. There is no effect on conjugated bilirubin and unconjugated bilirubin up to 40 mg/dL, hemoglobin up to 1 g/dL, and chyle up to 1000 FTU. However, do not use EDTA as it will affect the results. |
Detection devices : | Microplate reader, UV-visible spectrophotometer, colorimeter *The kit does not include 96-well plates or cuvettes. |
(1) Measurement of calcium concentration in control serums using this kit
Figure 1: Results of measurement of control serum with known calcium concentration.
(2) Measurement of serial dilution of calcium standard samples
Figure 2: Measurement results of calcium standard sample
Takuya Iwabuchi, Trace metal analysis and its pretreatment technology, Technical Information institute Co., Ltd. (ed.) p192-200 (2015)
Takuya Iwabuchi and Hiroko Suzuki, Development of diagnostic drugs/devices using cutting-edge biomarkers and regulatory compliance, Technical Information institute Co., Ltd. (ed.) (2015)
Masamitsu Kanai, Kanai's Manual of Clinical Laboratory Medicine, 35, p.201-203 (2005)
J. W. Ferguson, J. J. Richard, J. W. O’laughlin and C. V. Banks, “Simultaneous Spectrophotometric Determination of Calcium Chelate Color and Magnesium with Chlorophosphonazo-III “, Anal. Chem., 36, 796. (1964)
Michio ZENKI, Tomoko MASUTANI, and Takashi YOKOYAMA Repetitive Determination of Calcium Ion and Regeneration of a Chromogenic Reagent Using Chlorophosphonazo III and anIon Exchanger in a Circulatory Flow Injection System ANALYTICAL SCIENCES, VOL. 18, p1137-1140, OCTOBER 2002
In preparation
Contact Us
TEL：+81-19-681-8650
Telephone reception hours：Weekdays 9:00～17:00
（excluding weekends and holidays）
E-mail：morioka-sales@cellspect.com