Pathological changes in the spinal cords of rats were observed by hematoxylin-eosin (HE) staining. Nissl staining was performed to assess neuron survival, and ferric iron was visualized using Perls’ Prussian blue staining. Rats were anesthetized and sacrificed at 14 dpi. The spinal cord tissues from the injured epicenter were collected and fixed in 4% paraformaldehyde for 24–48 hours and then dehydrated for 12 hours using an automatic tissue dehydrator (ASP300S, Leica, Solms, Germany). After embedding in paraffin, samples were sliced by a microtome (RM2255, Leica) at 4 μm thickness. Before dewaxing and rehydration, sections were incubated for 2 hours at 60°C. After rehydration, sections were stained by HE Staining Kit (G1120, Solarbio, Beijing, China), Nissl staining solution (G1430, Solarbio) and Perls’ Prussian blue Staining Kit (G1422, Solarbio) following the manufacturer’s instructions. Finally, sections were dehydrated with gradient alcohol and made transparent with xylene, followed by mounting using neutral gums.
Perls prussian blue staining kit
Manufactured by Solarbio
Sourced in China
The Perls' Prussian blue Staining Kit is a laboratory tool used for the detection of iron deposits within biological samples. It provides a reliable and efficient method for visualizing ferric (Fe3+) ions through the formation of a distinctive blue-colored complex known as Prussian blue.
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5 protocols using perls prussian blue staining kit
1
Histopathological Analysis of Spinal Cord Injury in Rats
2
Quantifying Cellular Uptake of USPIO
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After 12 h incubation with varying concentrations of USPIO, cells were washed three times with PBS and intracellular iron oxide distribution was detected by Perl’s Prussian blue staining kit (Solarbio science & Technology, Beijing, China). Cells were fixed for 15 minutes in 4% paraformaldehyde, and then incubated for 25–30 minutes with 10% potassium ferrocyanide, rewashed twice with PBS, and counterstained with nuclear fast red for 10 minutes. Cells containing intracytoplasmic blue granules were defined to be Prussian blue staining positive (PB-positive). Cells labeled with varying concentrations of USPIO were trypsinized and washed 5 times with PBS, then detached, counted and resuspended in a 37% HCl solution. The iron content per cell was obtained by inductively coupled plasma mass spectroscopy (ICP-MS, MICI, Suisse). Three replicates at each concentration were measured for statistical analysis.
3
Tissue Iron Quantification via Perls Staining
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Brain slices underwent a thorough washing procedure with PBS prior to Perls Prussian Blue staining. The preparation of the Perls staining and the enhancement working solution adhered to the guidelines outlined in the Perls Prussian blue staining kit (G1428, Solarbio, China). Subsequent to completing the staining protocol as per the provided instructions, the brain slices underwent sequential dehydration in 70 % ethanol for 30 s, 80 % ethanol for 60 s, 90 % ethanol for 60 s, and absolute ethanol for 2 min. Following this, the slices were cleared in xylene for 5 min. Following the application of neutral resin for mounting, the slices underwent observation and microscopic imaging.
4
Intracellular Localization of Iron-Doped Carbon Dots
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B16 cells were cultured in RPMI-1640 containing 10% FBS and added with Fe-CDs and an equal volume of PBS. After 6 h, the cells were fixed with glutaraldehyde solution. Intracellular distribution of Fe-CDs was determined under transmission electron microscopy (80 kV, Philips EM208 or 120 kV, Tecnai12). The images were photographed at a magnification of × 50,000 or × 52,000, and scanned at 6 μm per pixel using an MRC-KZA scanner. High-resolution scanning electron microscope (FESEM, Merlin) demonstrated the intracellular distribution of Fe-CDs.
According to the manufacturer’s instructions, the Perl’s Prussian Blue Staining Kit (G1426, Solarbio, USA) was used to detect the distribution of intracellular iron ions. Briefly, the cells were fixed in 4% paraformaldehyde for 15 min, followed by incubation with 10% potassium ferrocyanide for 25–30 min, and then washed and restained with nuclear fast red for 10 min. The cells containing blue granules in the cytoplasm were considered positive for Prussian Blue staining.
According to the manufacturer’s instructions, the Perl’s Prussian Blue Staining Kit (G1426, Solarbio, USA) was used to detect the distribution of intracellular iron ions. Briefly, the cells were fixed in 4% paraformaldehyde for 15 min, followed by incubation with 10% potassium ferrocyanide for 25–30 min, and then washed and restained with nuclear fast red for 10 min. The cells containing blue granules in the cytoplasm were considered positive for Prussian Blue staining.
5
Intracellular Iron Oxide Localization
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The intracellular iron oxide distribution was detected using a Perls' Prussian blue staining kit (Beijing Solarbio Science & Technology, Beijing, China). The sections were fixed for 15 min in 4% paraformaldehyde and then incubated for 25-30 min with 10% potassium ferrocyanide, rewashed twice with PBS, and counterstained with Nuclear Fast Red for 10 min. Cells containing intracytoplasmic blue granules were defined to be Prussian blue (PB) staining-positive (PB-positive).
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