Histology

Histology

  • Mohamed A. A. Mahdy added an answer:
    How to identify adipocytes or fat containing cells in paraffin sections?
    Is there any way to staining or identify fat cells paraffin embedded muscle cross section? Oil red O requires frozen section.
    Mohamed Mahdy · Yamaguchi University
    @ Susan, thanks for the paper, but it need fixation in Osmium tetraoxide before paraffin embedding, I think Junxi has already paraffin sections not samples
  • Ahmed A Mohamedani added an answer:
    Without immunological markers, what cells can be accurately identified in a simple H&E section?
    In the literature, researchers often use antibodies against immune cell markers (CD4, CD11 and CD68) to quantify particular cells occurrence. What cells can be identified without such markers? For example, an atherosclerotic plaque is a hive of inflammatory cells. Other than the distinct foam cell structures, could intact macrophages be identified? Second question, are there any quick special stains that can identify leukocytes in tissue without performing IHC? Thank you!!
    Ahmed Mohamedani · University of Gezira
    I also agree withElizabith, but using quick stains is simple and give excellent morphlogy of most of the cells you are looking for. Such stains include diff quick, rapidiff and RAL. One can make his own quick stain to cut down expenses.
  • Paul Guyett asked a question:
    Does anyone have information on the histopath microscope?
    I am about to purchase a new upright bright field microscope to be used for histopathology. I am looking at Zeiss Axiolab and Olympus CX41 scopes. What are your experiences with the optics of these scopes? What scope do you recommend for brightfield microscopy? Optics, ergonomics, etc.
  • Eman Elkordy asked a question:
    What is the best way to prepare aflatoxin powder (solvent, dosage)?
    Going to give it to a rat.
  • Mohammad ali Jafari added an answer:
    Can anyone recommend a book containing high-quality histological images of polychaetes?
    I am currently interpreting histological slides of Sabellaria spinulosa in an attempt to determine their reproductive state and am struggling to find good images/explanations of the reproductive structures. Images in related journal articles are generally very small and only label major structures (e.g. eggs and sperm). I am really interested in the early developmental stages which are much harder to identify.
    Mohammad ali Jafari · Tarbiat Modares University
    Dear Bryony I have an application that is useful for identifying Polychaete worms. This software will ask you questions about the shape of your worm that eventually will lead to the genus or species. Due to high volume, I can not send it for you but according to the following references maybe you will find it. Wilson, R.S., Hutchings, P.A., Glasby, C.J., 2003. Ploychaetes, an interactive identification guide. Musium Victoria, Australian Museum, Museum and Art Gallery of the Northern Terriritory.
  • Salme Suhana added an answer:
    Does anyone know the correct way to cut the lung sample for asthmatic mice?
    I would like to see the effect of certain drug on the lung inflammation of ovalbumin induced mice however I am not sure how to cut the sample for H&E and periodic acid schiff (PAS) staining. Is it longitudinal or cross section? Can anyone help?
    Salme Suhana · Universiti Teknologi MARA
    Dear all, thank you for your kind response and suggestion. I've seen the website and it is very useful. I'll try to work out my sample and hopefully i can get a good view.
  • Roseanna Smither asked a question:
    Does anyone have a protocol for revealing BDA tracing in the brain with a Streptavidin Alexa Fluor conjugate?
    I am wanting to use BDA for neuronal tracing and rather than revealing this with DAB I would like to use a fluorescent tag, for viewing on the confocal microscope.
  • Does someone have the procedure to perform a toluidine blue coloration after the strips have been fixed?
    I have used Formaldehyde 10% to fixed rats organ after an experiment and I will make a post-fixation in bouin liquid before performing trimming and staining, but I don't know if it is compatible with toluidine blue stain. And I really don't know how to proceed, because I don't have any protocols to perform this stain.
    I normally use the Toluidine blue stain in plant tissues thus: we fix samples in FAA (50% ethanol+5% formaldehyde+10% glacial acetic acid, in water), then embed samples in paraffin to obtain think sections (using a microtome). After that, we apply 0.05% Toluidine blue (TBO) in PO4 buffer during 5–10 min. Finally, we remove paraffin to observe samples in the microscope. This process is very effective.
  • Ricardo Camarinho asked a question:
    What are the best cassette markers out there?
    I'm considering to buy a cassette marker for histological cassettes and slides. What are the best options? Laser or Ink? And what are the best machines out there, both affordable and with quality.
  • Wolfgang H. Muss added an answer:
    4% paraformaldehyde for fixing cells - 4 degrees or room temp?
    I see both used but no explanation. What's the truth here?
    Wolfgang Muss · Paracelsus Medical University Salzburg
    Dear Ahmed, you're welcome Best regards and wishes Wolfgang
  • Hans Snyder added an answer:
    Does anyone know a standard histological method for molluscan nervous system?
    Nervous system of molluscs and Histology.
    Hans Snyder · Worcester Polytechnic Institute
    Hello, I don't know much about molluscan but I do know histology. There are many stains for nerve fibers, astrocytes, myelin and nerve cells. I would first do H&E staining to assess the regions of interest. Then depending on what your looking for, do special stains. If you email me directly hans@histologistics.com with specifics about what parts of the nervous system you want, I will send you the staining protocols and tips to getting your project done. Our website http://histologistics.com/ has an abundance of pictures, protocols and other information about staining for all types of tissues. Let me know. Thank you
  • Bin Xu added an answer:
    What do you think of the criteria for determining the malignancy of "Oncocytic adrenocortical neoplasm"?
    What do you think of the criteria for determining the malignancy of "Oncocytic adrenocortical neoplasm" (Lin-Weiss-Bisceglia (LWB) system)? I refer in particular to venous invasion if this is the only criterion to indicate a malignant behavior. It could not be a "prolapse" into the veins as observed in the thyroid or kidney? Do you know other criteria to indicate the prognosis of these rare tumors?
  • Jesse M. S. Ellis added an answer:
    What can I do to improve the stability of my tissue sections when removing them from PBS-filled wells at the end of IHC?
    My 50um tissue samples stick to the bottom of my 96 well plates after completion of IHC. It is almost impossible to pipette them out for mounting without some kind of tearing. Any suggestions?
    Jesse Ellis · University of California, Los Angeles
    In both labs I've worked in we have used mesh bottomed inserts or trays. Tissue is transferred with a small paint brush (40µm) into the insert, and then the insert contains the tissue through the entire procedure, until the end. (You move the insert from tray to tray containing various reagents.) At the end the sections are floated in PBS onto gel-subbed slides.
  • Karen Nygard added an answer:
    Any tips on preventing cultivated epithelial cells coming loose from the scaffold during paraffin sections?
    I was wondering if there are any tips or tricks out there for insuring that cultivated epithelium does not detach from its collagen scaffold during sectioning prior to imaging. We have tried performing z stacks using confocal microscopy on whole mounts, but the current scaffolds are too think to get good stacks, so we need to revert to paraffin embedding to get tissue cross sections. Any ideas?
    Karen Nygard · The University of Western Ontario
    (pardon the typos. ..phone texting)
  • Claire M Dubois added an answer:
    How can I get western blot bands of hif-1a?
    I am trying to do western blotting of tumor samples for HIF-1a. The antibody is from neomarkers and I used a dilution of 1:1000. The protein samples from the tumor were isolated in RIPA buffer. I tried several times, but could not get the specified band. Any help would be highly appreciated.
    Claire Dubois · Université de Sherbrooke
    HI Vijayendra, If positive controls work (CoCL2 treated cells is indeed a good one), It might be because HIF-1a is degraded in your samples (half live less than 3 minutes). If it is indeed degraded, you can include proteasome inhibitors in your extraction (and perfusion if mouse tissues). If samples are not easy to get, you may want to use cabonic anhydrase 9. It is much more stable and in our hands a very robust marker of hypoxia. Good luck!
  • Sunil Londhe added an answer:
    Are there any histological techniques to identify sex of plant in seed embryo?
    Want to identify in seed stage it self with out any DNA markers
    Sunil Londhe · Shivaji University, Kolhapur
    In animal tissue we using MT- staining technique. Its result is cytoplasm of cells stain pink, nucleus stain dark red and other mussles stain blue. fron this you indentify the cytoplasm of cell and nucleus.
  • Sandra Mayr added an answer:
    Does anyone know an easy and reliable protocol for antigen-retrieval of FFPE-sections of heart muscle?
    I want to analyse formalin fixed and paraffin embedded tissue sections of human cardiomyocytes with immunofluorescence. Since formalin fixation disturbs protein antigens I need to retrieve them before staining with specific antibodies. Does anyone has experience which conditions fit best for cardiomyocytes (heat?, temperature? duration?) The protocol should ideally be easy to carry out - without the help of any special devices. Thanks in advance,
    Sandra Mayr · Fachhochschule Oberösterreich
    Thank you all very much!
  • Rhiannon Grant added an answer:
    Is anyone in the UK using the CLARITY method?
    I'd like to try the CLARITY method on some of my knock-in mice with fluorescent reporter molecules attached to their post synaptic proteins. It is impractical for me to set up the entire method when I don't know if it will kill the fluorescence, so it would be excellent if I could come and borrow someone's set up for a short period to test the method on my samples.
    Rhiannon Grant · The University of Edinburgh
    Hi Dirk and Michelle, I'm so sorry I didn't respond earlier - for some reason this thing hid your messages! I have tried some other clearing methods, including SeeDB and a urea based method whose name escapes me right now. I found SeeDB useful but we were concerned about the processing temperatures so we wanted to give Clarity a go. I tried the passive clearing and it did work, thank you for that tip Michelle!
  • Mohamed A. A. Mahdy added an answer:
    How can I prevent loss of tissue during H&E staining of cryostat section and what software can I use to quantify the size of polyps in the samples?
    I am trying to perform H&E staining on mouse guts (Swiss Roll). I have fixed the guts in NBF at room temperature overnight before preparing the Swiss Rolls. The cryosections that I made seem to be pretty good. However, when I started staining the tissue sections, they seem to be tearing off from the surface of the glass slide. I am using SuperFrost slides. Any suggestions on how I could prevent loss of tissue ? Also, once I am done with the H&E staining, what is the best way/software to quantify the size of the polyps in the samples? Any help would be greatly appreciated. Thanks
    Mohamed Mahdy · Yamaguchi University
    Thank you Breedge for the detailed protocol
  • Esraa Samir Tamim asked a question:
    Can anyone tell me the difference between a normal colon and colon cancer in rodents?
    There is a difference between normal and cancerous tissue of the colon of rodents after injection with 1,2- DMH for 15 weeks.
  • Vinay Lomash added an answer:
    Is it possible to do a golgi stain on freshly frozen sections?
    I would like to do a stain similar to the one described in: J Histochem Cytochem. Jun 2008; 56(6): 539–550. doi: 10.1369/jhc.2008.950246PMCID: PMC2386766A Rapid Method Combining Golgi and Nissl Staining to Study Neuronal Morphology and Cytoarchitecture I have freshly frozen sections from a mouse brain.
    Vinay Lomash · Jiwaji University
    see to this artical 'Quick Golgi method: Modified for high clarity and better neuronal anatomy' by patro et al 2013 if can work with few modification while fixation of slides in stain in coplin jar. i think this will be helpful and can give you one good publication also. try and validate this. best of luck its freely available on net http://nopr.niscair.res.in/handle/123456789/21074
  • Gaber Ramadan added an answer:
    Can anyone help with Hematoxylin and Eosin staining?
    I have just started staining my mouse hearts with H&E, although the staining works, my nuclei seems fainter than normal, and my eosin is extremely pink. I've tried decreasing the time for the Eosin from 4 minutes to 2 minutes, and increasing the time for Hematoxylin from 5 minutes to 7 minutes, but the results are the same. Should I try a longer differentiation time on the Eosin? I am using Mayer's H&E from Fisher to stain these tissue. The fixation time is one day in formaldehyde, and the sections are 6uM thick.
    Gaber Ramadan · United Arab Emirates University
    Hi Vinh Please, is it possible to mention the fixative you have used in fixing the mouse heart? Also, did you fix the whole heart or you cut it into small pieces? and what are the times you have used during dewaxing the mounted sections? Thank you
  • Leonardo Martins added an answer:
    Can anyone recommend a book or paper containing high-quality histological images and methods of sea anemone?
    About sea anemone, sea carpet (stichodactyla) or invertebrate.
    Leonardo Martins · Universidade Federal de São Paulo
    I agree with Aydin... just, advice you using free softwares for analyses and quantifications (like a ImageJ), as well.
  • Matthew Ibbs added an answer:
    Why H/E staining in cryostat sections breaks my tissue?
    I'm trying to do the staining H/E on cryostat sections of muscle (10 um). The tissue was fixed in 4% PFA and cryopreserved with 30% sucrose. When do the staining, samples are detached or disrupted. I let it air dry sections before fixing again with PFA for 5 min. Stain with Mayers H for 2 min, tap water, eosin 30s, tap water and EtOh 100% for 30s and xylene. I make the stain gently and horizontally. The problems begin on H and worsens on E and alcohols. Could someone suggest something?
  • Erin Dobrinen asked a question:
    Why the uneven Diff-Quick staining of lung homogenate?
    We use Diff-Quick (modified Wright-Giemsa) stain to find PC burden in our murine lung homogenates. We fix in methanol, stain in solution 1 for 3 minutes and solution 2 for 35 minutes. It usually gives us a crisp stain. Occasionally, however, we'll have a slide that is perfect, followed by a slide that is pale and unreadable. Now, both slides were in the same solution in the same rack. We've been careful not to touch the slide surface before the sample is applied by cytospin thinking that the oils from our hands are causing the problem. This hasn't made a difference. I thought I'd return to the original citation to see if we can find tips to optimize the stain. Many, many studies have been done comparing different PC staining techniques and describing each one in detail EXCEPT for Diff-Quik! Each paper I've read only refers to "we followed the manufacturer's protocol". We order the item through Fisher, and it takes a long time to arrive, seemingly because it moves through different suppliers. The original manufacturer has been taken over by other companies, more than once. It comes with no directions. As far as I can tell, there is no longer a manufacturer's protocol. I cannot find the item or item information on the websites that sold or sell the item (Baxter scientific, DADE Behring, Siemens). So, does anyone have an original protocol for using Diff-Quik to stain Pneumocystis??
  • Kuldip Gupta added an answer:
    Storage of tissue for immunohistochemistry?
    I am going to perform a mice experiment abroad and plan to harvest tissue (adipose tissue, liver) for subsequent histology and IHC. After one week I will return home. I know that fixing of tissue in 10% Formalin of more than 24 hrs may affect further IHC. I am grateful for advice about the best storage/transport medium after fixing?
    Kuldip Gupta · Guru Angad Dev Veterinary and Animal Sciences University
    I agree with all the above answers. The prolonged fixation with formalin can lead to cross linking of polymers and it may interfere with IHC staining. Its over experience that if you face such problem then you can use a combination of HIER protocols. In our laboratory we use following protocol Exposure to Dewaxing solution at 70 C for 10 minutes Exposure to Cirtrate buffer 95 C for 10 minutes Followed by exposure to TRIS -EDTA buffer for 10 minutes. The above protocol has given very good results in studying the expression of angiogenic & lymphangiogenic markers, heat shock protein, Cox-2 and BCRP and MDR in tissue paraffin embeded tissue sections of canine mammary tumors( un published results) even when the tissues were kept in 10 % NBF for around7-10 days.
  • Jessica Laurence added an answer:
    Has anyone sectioned tissues preserved in RNAlater?
    I'm trying to slice unfixed bird brains that were preserved at -20°C in RNAlater. It seems that the tissues are not frozen properly when I'm trying to cut them in a cryostat, probably because of the RNAlater (the tissue was frozen in tissue-tek on dry ice before). The tissue unravels inside the mold when I try to slice it. I want to perform an in situ hybridization on these brains. Anybody have a solution?
    Jessica Laurence · University of Adelaide
    We tried a technique similar to Marilyn recently. We had good results by putting the tissue in RNA later in the fridge overnight (mouse organs) in eppendorf tubes. The next day we bisected the tissues, keeping half in RNA later (stored at -80oC for RNA) and fixing the other half. We washed out the PBS with 2x washes for 1h each, then fixed in 10% neutral buffered formalin overnight and took the tissues through the normal fixation process. The tissues stained well. I wouldn't advise freezing the tissues in RNA later before fixation as this may distort the tissue.
  • Shingo Takano added an answer:
    What type of cancer reveals glomeruloid vessels when observing the histology?
    Glomeruloid vessels are a hallmark of Glioblastoma histology but are there any other cases of cancer?
    Shingo Takano · University of Tsukuba
    Glomeruloid vessel is seen on glioboastoma section, called microvascular proliferation.
  • Is there a way to speed up release of coverglass in xylenes from DAB-stained/DPX-mounted sections?
    I have some slides I did a DAB stain on from early last summer that I'd like to counterstain with methyl green. Everything was DAB stained, dehydrated (50% EtOH, 75% EtOH, 95% EtOH, 2x 100% EtOH, 2x xylenes) and mounted with DPX mounting medium. My mentor suggested leaving them in xylenes until the cover glass slides right off so that I can rehydrate and counterstain my sections. The only problem is, these slides were mounted in June last year and the sections have been in xylenes now for 96 hr+ and the cover glass won't come off. Any tips? I can get a DAB stain and methyl green counterstain to work just fine (see attached), so I'd like to get my older sections to look like my newer work.
    Ayman EL-Kenawy · Taif University. Sadat city Univeristy,
    Dear Aaron Good job.

About Histology

The study of the microscopic anatomy of cells and tissues of plants and animals, including tissue fixing, fixation and staining.

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