scholarly journals A technique for retrieving antigens in formalin-fixed, routinely acid-decalcified, celloidin-embedded human temporal bone sections for immunohistochemistry.

1992 ◽  
Vol 40 (6) ◽  
pp. 787-792 ◽  
Author(s):  
S R Shi ◽  
C Coté ◽  
K L Kalra ◽  
C R Taylor ◽  
A K Tandon
Keyword(s):  
Temporal Bone ◽  
Room Temperature ◽  
Antigen Retrieval ◽  
Positive Staining ◽  
Retrieval Technique ◽  
Published Report ◽  
Human Temporal Bone ◽  
Antigen Loss ◽  
Formalin Fixed ◽  
Fixed Acid

The application of immunohistochemistry to routinely decalcified, celloidin-embedded human temporal bone sections has been hampered because of antigen loss during processing of the specimens. To our knowledge, there has been no published report to date describing immunohistochemical staining of such tissues suitable for examination by light microscopy. Here we report a novel antigen retrieval technique which can be successfully used to stain a variety of antigens in routinely formalin-fixed, trichloroacetic acid-decalcified, celloidin-embedded human temporal bone sections. The new procedure reported here for decalcified human temporal bone tissues simply requires immersing slides for 30 min at room temperature in an antigen retrieval solution. A total of 60 decalcified, celloidin-embedded human temporal bone tissues were tested with monoclonal antibodies (MAb) to 15 different antigens. Of these, 12 MAb showed definite positive staining, while three were negative. This technique may prove very useful in studying the expression of various antigens by immunohistochemistry in formalin-fixed, acid-decalcified, celloidin-embedded tissues.

scholarly journals Protease Antigen Recovery Decreases the Specificity of Bromodeoxyuridine Detection In Formalin-fixed Tissue

1997 ◽  
Vol 45 (8) ◽  
pp. 1165-1170 ◽  
Author(s):  
Philip M. Bak ◽  
Ralph J. Panos
Keyword(s):  
Cellular Proliferation ◽  
Enzymatic Digestion ◽  
Antigen Retrieval ◽  
Alveolar Type ◽  
Positive Staining ◽  
Specific Staining ◽  
Fixed Tissue ◽  
Formalin Fixed Tissue ◽  
Protease Digestion ◽  
Formalin Fixed

Incorporation of halogenated nucleotide analogues is often used to assess DNA synthesis and to quantitate cellular proliferation. Multiple antibodies have been developed to bromodeoxyuridine (BrdUrd) and it is the most frequently utilized substrate. Because the immunodetection of incorporated BrdUrd requires DNA denaturation or nuclease digestion, most of these antibodies are not reactive in tissues or cells fixed with crosslinking agents. Antigen retrieval techniques utilizing protease digestion restore BrdUrd antigenicity and permit the detection of BrdUrd in formalin-fixed tissue. However, during the development of a double label immunohistochemical protocol to quantitate proliferating alveolar Type II cells, we noted nucleus-specific staining in lung sections from animals that had not received BrdUrd. Therefore, we systematically analyzed the specificity of the immunohistochemical detection of incorporated BrdUrd in formalin-fixed tissue after protease digestion. Enzymatic antigen recovery diminished the specificity of the BrdUrd reaction product and caused false-positive staining with the BU-1, B44, and BR3 monoclonal antibodies. Staining was less prominent with Bu20a but was more specific. Protease antigen recovery may decrease the specificity of BrdUrd immunodetection. Appropriate controls are required when enzymatic digestion is used to detect incorporated BrdUrd in formalin-fixed tissue. The type and duration of fixation, antibody to BrdUrd, protease, and tissue may affect the specificity of the staining pattern. (J Histochem Cytochem 45:1165–1170, 1997)

Standardization of Immunohistochemistry Based on Antigen Retrieval Technique for Routine Formalin-fixed Tissue Sections

1998 ◽  
Vol 6 (2) ◽  
pp. 89-96 ◽  
Author(s):  
Shan-Rong Shi ◽  
Richard J. Cote ◽  
Benjaporn Chaiwun ◽  
Lillian L. Young ◽  
Yan Shi ◽  
...  
Keyword(s):  
Antigen Retrieval ◽  
Fixed Tissue ◽  
Formalin Fixed Tissue ◽  
Tissue Sections ◽  
Retrieval Technique ◽  
Formalin Fixed

Immunohistochemistry of Lymphocytes and Macrophages in Human Celloidin-Embedded Temporal Bone Sections with Acute Otitis Media

1997 ◽  
Vol 106 (8) ◽  
pp. 662-668 ◽  
Author(s):  
Tetsuya Ganbo ◽  
Isamu Sando ◽  
Chiaki Suzuki ◽  
Carey D. Balaban ◽  
Masaharu Sudo
Keyword(s):  
Temporal Bone ◽  
Otitis Media ◽  
Acute Otitis Media ◽  
Microwave Treatment ◽  
Antigen Retrieval ◽  
Suppurative Otitis Media ◽  
Human Temporal Bone ◽  
Air Cells ◽  
Immunohistochemical Analyses ◽  
Mastoid Air Cells

Immunohistochemical analyses were used to investigate the distribution of lymphocytes and macrophages in routine human temporal bone sections obtained from a subject with acute suppurative otitis media. Primary antibodies specific for human CD3 and CD43 (T-lymphocytes), CD20 (B-lymphocytes), CD45 (leukocyte common antigen), and CD68 (macrophages) were used. As a pretreatment, the sections were soaked in antigen retrieval solution (saturated sodium hydroxide-methanol solution in methanol at a ratio of 1:3). A second antigen retrieval procedure (microwave treatment in 1 % zinc sulfate) was also employed for identifying CD3-positive cells. Then the avidin-biotin-peroxidase complex technique was performed. Positive reactions to all antibodies but anti-CD68 were observed in the mucosa of the eustachian tube, tympanic cavity, and mastoid air cells. Particularly, cells positive to anti-CD3 or anti-CD43 were making a diffuse invasion upon the lamina propria. CD68-positive cells were scattered only in the effusion of mastoid air cells. These results suggest that the retrospective immunohistochemical study of archival temporal bone sections is a promising approach to investigate the pathogenesis of otitis media.

scholarly journals Comprehensive Evaluation of PAXgene Fixation on Oral Cancer Tissues Using Routine Histology, Immunohistochemistry, and FTIR Microspectroscopy

Biomolecules ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 889
Author(s):  
Pooja Lahiri ◽  
Suranjana Mukherjee ◽  
Biswajoy Ghosh ◽  
Debnath Das ◽  
Basudev Lahiri ◽  
...  
Keyword(s):  
Comprehensive Evaluation ◽  
Antigen Retrieval ◽  
Tissue Fixation ◽  
Ftir Microspectroscopy ◽  
Protein Secondary Structures ◽  
Associated Proteins ◽  
Cancer Tissues ◽  
Formalin Fixed ◽  
Fixation System

The choice of tissue fixation is critical for preserving the morphology and biochemical information of tissues. Fragile oral tissues with lower tensile strength are challenging to process for histological applications as they are prone to processing damage, such as tissue tear, wrinkling, and tissue fall-off from slides. This leads to loss of morphological information and unnecessary delay in experimentation. In this study, we have characterized the new PAXgene tissue fixation system on oral buccal mucosal tissue of cancerous and normal pathology for routine histological and immunohistochemical applications. We aimed to minimize the processing damage of tissues and improve the quality of histological experiments. We also examined the preservation of biomolecules by PAXgene fixation using FTIR microspectroscopy. Our results demonstrate that the PAXgene-fixed tissues showed significantly less tissue fall-off from slides. Hematoxylin and Eosin staining showed comparable morphology between formalin-fixed and PAXgene-fixed tissues. Good quality and slightly superior immunostaining for cancer-associated proteins p53 and CK5/6 were observed in PAXgene-fixed tissues without antigen retrieval than formalin-fixed tissues. Further, FTIR measurements revealed superior preservation of glycogen, fatty acids, and amide III protein secondary structures in PAXgene-fixed tissues. Overall, we present the first comprehensive evaluation of the PAXgene tissue fixation system in oral tissues. This study concludes that the PAXgene tissue fixation system can be applied to oral tissues to perform diagnostic molecular pathology experiments without compromising the quality of the morphology or biochemistry of biomolecules.

scholarly journals Antigen retrieval technique utilizing citrate buffer or urea solution for immunohistochemical demonstration of androgen receptor in formalin-fixed paraffin sections.

1993 ◽  
Vol 41 (11) ◽  
pp. 1599-1604 ◽  
Author(s):  
S R Shi ◽  
B Chaiwun ◽  
L Young ◽  
R J Cote ◽  
C R Taylor
Keyword(s):  
Androgen Receptor ◽  
Enzymatic Digestion ◽  
Antigen Retrieval ◽  
Urea Solution ◽  
Paraffin Sections ◽  
Buffer Solution ◽  
Solution Ph ◽  
Citrate Buffer ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed

We developed a staining protocol for demonstration of androgen receptor (AR) in formalin-fixed, paraffin-embedded tissue sections. The method is based on the antigen retrieval microwave (MW) heating technique. Results are compared with different types of enzyme digestion pre-treatments. The strongest immunostaining signal and clearest background were obtained by MW heating of dewaxed paraffin sections in 5% urea or citrate buffer solution (pH 6); pure distilled water gave less consistent results. Enzymatic digestion with pepsin (0.05% in 2 N HCl) for 30 min at room temperature, or trypsin followed by pronase, or pronase digestion alone, also produced enhanced staining of AR in some cases, but there was more nonspecific background, and specific reactivity was less intense. The antigen retrieval MW method can be used to demonstrate AR epitope in prostate tissue after fixation in formalin for as long as 7 days. AR immunolocalization was also compared in frozen and paraffin sections processed from the same specimen of prostate carcinoma tissue and was found to be qualitatively and quantitatively similar. This study also provided new information concerning the basic principles of the antigen retrieval MW method that may be helpful in further development of this technique.

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