Hydroxyproline Assay试剂盒(Colorimetric)

• Biochemical assay

Lab

Biochemical assay - Hydroxyproline Assay Kit (Colorimetric) (AB222941)

Diagram showing the principles of the Hydroxyproline assay method.

• Biochemical assay

PubMed

Biochemical assay - Hydroxyproline Assay Kit (Colorimetric) (AB222941)

Patrawalla et al used Hydroxyproline Assay Kit ab222941 to investigate hydroxyproline content in different human-derived tissue ECM.

Hydroxyproline assay kit was purchased from Abcam (Cat# ab222941/K226) (Cambridge, UK).

To quantify the collagen content in tissue ECM solutions, the amount of hydroxyproline (Hyp) was measured using a hydroxyproline assay kit. Briefly, tissue ECM solutions (N = 3/group) were mixed with equal volumes of 10N NaOH and hydrolyzed at 120 °C for 1 h. Following this, the mixture was cooled on ice, and neutralized with 10N HCl. The mixture was then centrifuged at 8000× g for 5 min and the supernatant was collected in a new tube. The sample hydrolysate was then evaporated by incubating the plate at 65 °C after transferring 10 µL aliquots from each tube to a 96-well plate. Further, 100 µL of the oxidation reagent mix was added to each well, and the plate was incubated at room temperature for 20 min. Next, 50 µL of developer solution was added to each well, and the plate was incubated at 37 °C for 5 min. The DMAB concentrate solution was then added to each well, and the plate was incubated at 65 °C for 45 min. Following this, the absorbance was measured in a M2 Spectramax plate reader (Molecular Devices) at 560 nm. A standard curve was obtained by measuring the absorbance of known Hyp concentrations ranging from 0–0.1 µg/µL. The Hyp content in the tissue ECMs was calculated by using Equation (1), where B is the amount of hydrolyzed Hyp (µg) calculated from the standard curve, V is the sample volume (µL) added to the well, and D is the dilution factor of the sample to fit in the range of the standard curve.

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Functional Studies - Hydroxyproline Assay Kit (Colorimetric) (AB222941)

Typical Hydroxyproline standard calibration curve.

• FuncS

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Functional Studies - Hydroxyproline Assay Kit (Colorimetric) (AB222941)

• Biochemical assay

PubMed

Biochemical assay - Hydroxyproline Assay Kit (Colorimetric) (AB222941)

Zhu et al used Hydroxyproline Assay Kit ab222941 to investigate explore MQZJFD as an innovative therapeutic agent for pulmonary fibrosis using bleomycin (BLM)-treated rats.

Effects of MQZJFD on the histopathological examinations and collagen examinations on the lung tissues of BLM-treated rats. The hydroxyproline content in lung tissues of BLM-treated rats.

Determination of the hydroxyproline content in lung tissues of BLM-treated rats

The hydroxyproline level in the lung tissues of rats were measured using the hydroxyproline assay kit obtained from Abcam (ab222941). Briefly, lung issues (100 mg) were homogenized in 100 µL of dH2O. Then, 100 µL of sample homogenate was transferred to a new screw-capped tube, and added 100 µL of NaOH (10 M) to the homogenate and incubated at 120 °C for 1 h. All samples were cooled down on ice before adding 100 µL of HCl. Finally, the tubes were centrifuged at 10,000 x g for 5 min. Hydroxyproline level in the supernatant was detected as the manufacturer’s instructions.

• Biochemical assay

PubMed

Biochemical assay - Hydroxyproline Assay Kit (Colorimetric) (AB222941)

Zou et al used Hydroxyproline Assay Kit ab222941 to investigate pulmonary induced fibrosis in mice.

Inhibition of SIK2 attenuates bleomycin (BLM)-induced pulmonary fibrosis in mice. Male BALB/C mice were anesthetized and received intranasal instillation of BLM (5 mg/kg) or its saline vehicle, followed by receiving administration of ARN-3236 (10, 30 mg/kg), CREB inhibitor 666-15 (10 mg/kg) or their vehicle. Lung tissues were isolated at day 28 for analysis. The content of hydroxyproline in lung tissues. Semiquantitative analysis of lung tissues by H&E staining.

Hydroxyproline assay

The pulmonary hydroxyproline content were measured using a previous reported method [17]. In brief, lung tissue (30 mg) was homogenized in 10 N concentrated NaOH (1 mL), and the mixtures were incubated for 60 min at 95 °C. After cooling to room temperature, the hydrolysates were neutralized to pH 6.0–6.8 with 10 N concentrated HCl, following by centrifugation at 10,000×g for 5 min at room temperature. The supernatant was collected, and the hydroxyproline content was evaluated with a hydroxyproline assay kit (Abcam, ab222941) following the manufacturer’s instructions. The results were expressed as µg hydroxyproline/ mg lung tissue.

• Biochemical assay

PubMed

Biochemical assay - Hydroxyproline Assay Kit (Colorimetric) (AB222941)

Awwad et al used Hydroxyproline Assay Kit ab222941 to examine cardiac remodeling in mice following transverse aortic constriction (TAC) in the presence or absence of proliferating cancer cells.

Tumor growth suppresses fibrosis in TAC-operated female mice. Representative paraffin-embedded heart sections of TAC-operated, PyMT-bearing and non-cancer-bearing female mice stained with Masson's trichrome to visualize fibrosis. Scale bar : 500 μm (top) and 50 μm (bottom). Quantification of the level of fibrosis (%) by Masson's trichrome staining in TAC-operated, PyMT-bearing (n = 4) and non-cancer-bearing female mice (n = 4), at least 5 random fields were chosen per mouse. Fibrosis hallmark gene marker Col1a1 in TAC-operated, PyMT-bearing (n = 8) and non-cancer-bearing (n = 8) female mice, measured using qRT-PCR, normalized to the GAPDH housekeeping gene. Hydroxyproline levels in the heart lysates of TAC-operated, PyMT-bearing (n = 4) and non-cancer-bearing (n = 4) female mice. The results are presented as mean ± SEM, Student's t-test. * p < 0.05, ** p < 0.01, *** p < 0.001. Each dot represents one mouse.

The hydroxyproline content was assessed as an index for collagen and fibrosis content in cardiac cell lysates using a colorimetric hydroxyproline assay kit (Abcam, USA, ab222941). The assay was performed according to the manufacturer's instructions.

• Biochemical assay

PubMed

Biochemical assay - Hydroxyproline Assay Kit (Colorimetric) (AB222941)

Park et al used Hydroxyproline Assay Kit ab222941 to investigate how L-carnosine attenuates bleomycin-induced oxidative stress in the pathogenesis of pulmonary fibrosis.

L-carnosine downregulated collagen deposition in bleomycin-induced fibrosis. It was pictured various regions including whole lung (top), proximal (middle), and distal (bottom). The lung tissue was stained using M/T staining. Pathological lesions were indicated with black arrows. Ashcroft score of each group. Hydroxyproline contents in tissue of each group (n = 4/group).

For measurement of hydroxyproline in lung tissue, it was performed using hydroxyproline colorimetric assay kit (ab222941, Abcam). For sample preparation, the lung tissues were reacted with 10 N NaOH for 1 h. Additionally, then, it was neutralized with 10 N HCl. Afterward, we followed the manufacturer’s protocol. Briefly, oxidation mixture was added to supernatant and reacted for 20 min. The developer and DAMB were added per well, and reacted at 65 °C for 45 min. Finally, it was measured at 560 nm by a microplate reader (BioTek, Winooski, VT, USA).

• Biochemical assay

PubMed

Biochemical assay - Hydroxyproline Assay Kit (Colorimetric) (AB222941)

Hong et al used Hydroxyproline Assay Kit ab222941 to investigate kidney health in mice treated with unilateral ureteral obstruction (UUO).

Kidney hydroxyproline content (sham, n=8; UUO, n=12).

Hydroxyproline content

Kidney hydroxyproline content was determined using a Hydroxyproline Assay Kit (Colorimetric) (ab222941, Abcam) and normalized to total protein measured by Bradford 1Ч Dye Reagent.

• Biochemical assay

PubMed

Biochemical assay - Hydroxyproline Assay Kit (Colorimetric) (AB222941)

Kumar et al used Hydroxyproline Assay Kit ab222941 to evaluate the therapeutic efficacy of MDB5 loaded micelles in common bile duct ligation (CBDL) induced liver fibrosis.

GDC-0449 and MDB5 loaded micelles inhibit progression of CBDL-induced liver fibrosis. Hydroxyproline content. IHC staining for protein expression of Ki67 (Objective 10X, inset objective 40 X).

Hydroxyproline content of liver samples was determined using the hydroxyproline assay kit (# K226-100, Biovision [Biovision assay kits are now manufactured by Abcam]) according to the manufacturer's instructions. Briefly, liver tissues (50 mg) were homogenized using a bead mill homogenizer and then hydrolyzed with 10 N HCl. Next, 10N NaOH was added to neutralize, and the sample hydrolysate was dried and oxidized with an oxidation reaction mix. The bright-colored chromophore was developed with proprietary acidic developer and absorbance was measured at 560 nm. Hydroxyproline concentration was calculated from a standard curve prepared with pure hydroxyproline and expressed as mg hydroxyproline per gram of liver tissue.

• Biochemical assay

PubMed

Biochemical assay - Hydroxyproline Assay Kit (Colorimetric) (AB222941)

Zhang et al used Hydroxyproline Assay Kit ab222941 to investigate progression of nonalcoholic steatohepatitis (NASH).

Representative images of Masson’s trichrome (MTS) and Sirius red stained liver sections of 16-week-old mice with indicated genotypes after feeding with CD-HFD for 8 weeks. Graph depicting percentage of MTS+ and Sirius red+ area on liver sections of indicated genotypes. n = 6 mice for each genotype. Liver hydroxyproline in livers of 16-week-old mice with indicated genotypes after feeding with CD-HFD for 8 weeks. n = 6 mice for each genotype. Each dot represents an individual mouse.

Hydroxyproline measurement : Liver tissue was homogenized in distilled water. 100 μl lysate was hydrolyzed with 100 μl 10 N NaOH at 120 oC for 1 h. Lysate was cooled on ice and neutralized with 1 N HCl. Hydroxyproline in supernatant was measured with Hydroxyproline assay kit (Abcam, Cat. ab222941), according to the manufacturer’s instruction.

• Schematic Diagram

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Schematic Diagram - Hydroxyproline Assay Kit (Colorimetric) (AB222941)

Representative image of Hydroxyproline Assay Kit (Colorimetric) ab222941

Components shown from left to right :

- Chloramine T Concentrate

- Hydroxyproline Standard

- Developer Solution I

- DMAB Concentrate

- Oxidation Buffer

- AlumaSeal 96 Film

Note : The vial labels shown in this image use generic names for illustrative purposes only and may not exactly match the specific component names included in the kit.

Note : Colors of solutions in image may not precisely match the shade of colors in the actual kit.