Tissues were fixed with 10% formalin, and stained with hematoxylin and eosin. Whole bone marrow cells were also prepared by cytospin and stained using Kwik-Diff (Fisher Scientific).
Kwik diff
Manufactured by Thermo Fisher Scientific
Sourced in United States
Kwik-Diff is a rapid staining system designed for the differentiation and identification of cellular components in biological samples. It provides a quick and consistent staining process for various types of cells, including blood cells, cytological preparations, and microbiological specimens. The Kwik-Diff system utilizes a series of staining solutions to highlight the morphological features of the cells, enabling efficient analysis and diagnosis.
Automatically generated - may contain errors
Lab products found in correlation
24 gauge angiocatheter, by BD (2 mentions)
Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Superfrost plus slide, by Thermo Fisher Scientific (2 mentions)
Bleomycin, by Cayman Chemical (1 mentions)
Facscalibur, by BD (1 mentions)
Upright light microscope, by Nikon (1 mentions)
Varian cary 50, by Agilent Technologies (1 mentions)
Pannoramic 250 flash, by 3DHISTECH (1 mentions)
Axiooberver, by Zeiss (1 mentions)
Phosphate buffered saline (pbs), by Merck Group (1 mentions)
Ez single cytofunnel, by Thermo Fisher Scientific (1 mentions)
Calcein am, by Thermo Fisher Scientific (1 mentions)
Eclipse e600, by Nikon (1 mentions)
Ly6c percp cy5, by BioLegend (1 mentions)
Multisizer, by Beckman Coulter (1 mentions)
Viability dye efluor 780, by Thermo Fisher Scientific (1 mentions)
Cd206 pe, by BioLegend (1 mentions)
F4 80 pe cy5, by BioLegend (1 mentions)
Cd11c af700, by BioLegend (1 mentions)
Trustain fc block, by BioLegend (1 mentions)
15 protocols using kwik diff
1
Formalin Fixation and Hematoxylin-Eosin Staining
2
Bleomycin-Induced Lung Injury in Mice
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Mice were anesthetized with ketamine/xylazine (ketamine (125 mg/kg; Sanofi Winthrop Pharmaceuticals, New York, NY), xylazine (12.5 mg/kg; Phoenix Scientific, St. Joseph, MO) and injected intratracheally using a 24-gauge angiocatheter with 100μl sterile PBS or bleomycin (Cayman Chemicals, Arbor, MI, 0.75 mg/kg dissolved in PBS). Four days later, animals were sacrificed and bronchoalveolar lavage fluid (BALF) was collected by instilling 1 mL chilled PBS into the lungs via a 24-gauge angiocatheter (Becton Dickinson) and collected. The procedure was repeated 5X each with fresh PBS. The BALF was then centrifuged for 5 min at 300 g. as analyzed for total cell count and leukocyte differential. Cells were characterized morphologically and counted by adhering them to glass slides using the Cytospin system (Shandon, Southern Sewickley, PA) and then staining them by Kwik-Diff (Fisher Scientific, Boston MA). The adhered stained cells were air-dried and counted in a blinded manner under a light microscope.
3
Neutrophil Quantification in Mouse BALF
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Three days after bleomycin and 24h after HCL and LPS exposure, mice were anesthetized with an intraperitoneal injection of ketamine (125 mg/kg; Sanofi Winthrop Pharmaceuticals, New York, NY) and xylazine (12.5 mg/kg; Phoenix Scientific, St. Joseph, MO). The trachea was cannulated using a (24 gauge angiocatheter (Becton Dickinson), and BALF collected as described above. Filtered BALF and red blood cell–lysed blood cells were resuspended in FACS buffer (Mouse PBS, 2% fetal calf serum, 2 mM EDTA). Cells were stained with the two monoclonal specific monoclonal antibodies against neutrophils Anti-CD11B-PE and Ly6G-FITC (BD Bioscience, Woburn, MA) (20 min, 4 °C on ice). Cells were then washed and resuspended in 50 μl FACS buffer. All studies were performed on a FACS Calibur (Becton Dickinson, San Jose, CA), and data were analyzed with FlowJo software (Tree Star, Ashland, OR). To confirm the presence of neutrophils within the different flow cytometry samples, we separately characterized a sample from each group by adhering the cells to glass slides using the Cytospin system (Shandon, Southern Sewickley, PA) and then staining them by Kwik-Diff (Fisher Scientific, Boston MA). The adhered stained cells were air-dried and counted in a blinded manner under a light microscope.
4
Bronchoalveolar Lavage Cell Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
BAL was done at the indicated times after the last OVA-challenge in anesthetized mice by intubating and flushing the airways 3 times with PBS for a total volume of 1 ml. The total number of cells in BAL was enumerated in a Neubauer hemocytometer and differential cell count was done by morphological examination of >300 cells on Kwik-Diff (Thermo Fisher Scientific Inc., Pittsburgh, PA)-stained cytospin slides. Eosinophil, lymphocyte, neutrophil and macrophage counts were calculated by multiplying the total BAL cell count with cell percentages.
5
Neutrophil Apoptosis Quantification Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Peripheral blood neutrophils were purified from healthy volunteers by dextran sedimentation and Percoll gradients as previously described [89 (link)]. Neutrophils were cultured at 37 ᵒC / 5% CO2 for 6 hours with either DMSO (negative control) or the compounds stated, in duplicate. Cytospins were produced with a cytocentrifuge at 300 rpm for 3 minutes for each condition. Cytospin slides were fixed with methanol then stained with Kwik-Diff (Thermo Scientific). Using an upright light microscope (Nikon) at 100x magnification, neutrophils were classified as either apoptotic or non-apoptotic based on the morphology of the nuclei. 1000 Neutrophils were counted per condition (500 per cytospin) to calculate percentage neutrophil apoptosis.
6
Bronchoalveolar Lavage Analysis Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
BAL fluids were centrifuged at 1500 rpm for 5 min at room temperature (Hettich GmbH). Supernatants were aliquoted and stored at −80 °C (Brunswick, Eppendorf). Pellets were resuspended in PBS (Merck‐Sigma), cells were counted and cytospun at 600 rpm for 5 min with ≈100 000 cells per slide (Hettich GmbH). Slides were then fixed in 100% ice‐cold methanol for 10 min, air‐dried and stored at −20 °C. Differential cell staining was performed using Kwik‐Diff (Shandon, ThermoFisherScientific) following the provider's instructions. The number of neutrophils, eosinophils, mono‐ and multinucleated macrophages, and lymphocytes were assessed using optical microscopy (AxioOberver, Zeiss). Colorimetric bright‐field images (Figure 5c ; Figure S1a , Supporting Information) were generated with a slide scanner (Pannoramic 250 Flash, 3DHistech Ltd). The protein content in BAL supernatants was evaluated using a BCA assay (Pierce BCA Protein Assay Kit, ThermoFisherScientific) following provider's instructions. Absorbance values were recorded at 562 nm using a spectrophotometer (Varian Cary 50, Agilent) and concentration values were calculated using a standard curve.
7
Bronchoalveolar Lavage Fluid Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Immediately after sacrifice BAL fluid was collected for assessment of the inflammatory responses in the lungs. Briefly, the trachea was exposed and cannulated allowing the lungs to be lavaged with 500 μl ice-cold sterile PBS. The lungs were gently aspirated and the process repeated a further two times (1.5 ml total). BAL fluid was centrifuged at 1500×g for 10mins at 4 °C and the supernatant from the first lavage was collected for subsequent cytokine enzyme-linked immunosorbent assay (ELISA) analysis. BAL Cells were counted using a haemocytometer and results expressed as total BAL cell counts (Additional file 2 ). Cells were adjusted to a concentration of 1 × 106 cells/ml, centrifuged using EZ single cytofunnels (Thermo Scientific, cat# 97200125) onto Superfrost™ Plus slides (Menzeil-Gläser Superfrost Plus, Thermo Scientific, cat# 10149870). Cells were then stained for differential immune cell counts using a Wright-Giemsa stain (Kwik Diff, Thermo Scientific, cat# 9990700). Slides were dried overnight before mounting and counting macrophages, lymphocytes, neutrophils and eosinophils across a minimum of 300 cells per sample. Cells were counted for each animal over three different slides.
8
Quantifying Airway Inflammation via BAL
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
To evaluate airway inflammation, bronchoalveolar lavage (BAL) was carried out via tracheostomy. A cannula was inserted into the trachea and then flushed with a total of 1 ml of PBS. The leukocyte cells in the BAL fluid were counted with a hemocytometer (Neubauer chamber). The BAL fluid was cytocentrifuged (Cytospin-4, Shandon Instruments, UK) and then stained with Kwik-Diff (Thermo Fisher Scientific Inc., Pittsburg, PA). Macrophages, eosinophils, neutrophils and lymphocytes were then enumerated based on morphological examination. A total of at least 300 cells per sample were counted.
9
Transwell Assay for Cell Migration and Invasion
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Tumor cells were harvested in serum-free media and seeded at 0.5–1 × 105 cells per well onto 6.5 mm × 8 μm pore size Transwell filters (3422; Corning), either uncoated for migration or Matrigel-coated (354234; Corning) for Matrigel invasion. For LP9 invasion, LP9 cells were seeded at 6 × 104 cells per well and grown to a confluent monolayer on thin type I collagen-coated (10 μg/cm2) Transwell filters for 24 h. OvCa cells were harvested, stained with calcein-AM (C1430; Invitrogen), and seeded at 1 × 105 cells per well onto LP9 and collagen-coated Transwell filters. Bottom chambers were filled with 400 μl of serum-containing media to allow cells to invade toward a serum gradient. After 24 h of tumor cell invasion, the top of the Transwell filters were wiped with a cotton swab and the bottom side was either stained with Kwik-Diff (9990705; Thermo Fisher Scientific) according to manufacturer’s instructions (for migration and Matrigel invasion assays) or fixed with PFA for 15 min, and fluorescently labeled invaded cells were visualized by EVOS FL Auto Cell Imaging (LP9 invasion assays). All migrated and invaded cells per Transwell were counted using the multipoint tool on ImageJ; fluorescence intensity of labeled invaded cells in LP9 invasion assays was measured by ImageJ.
10
Murine Bronchoalveolar Lavage and Cytology
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Bronchioalveolar lavage (BAL) was performed by instilling and collecting 1 ml of sterile saline into the lungs of euthanized mice. BAL cells were pelleted and counted and 5x104 cells in 200 μl PBS were centrifuged in a Cytospin. Preparations were air dried overnight and then and stained using Kwik-Diff (Thermo Scientific). Pictures were visualized in a Nikon Eclipse E600 microscope.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!