HistoChoice® as an alternative to formalin fixation of undecalcified bone specimens

We compared histochemical and immunohistochemical staining as well as fluorochrome labeling in murine bone specimens that were fixed with 10% neutral buffered formalin to those fixed with HistoChoice®. We showed that sections from undecalcified tibiae fixed for 4 h in HistoChoice® resulted in enhanced toluidine blue and Von Kossa histochemical staining compared to formalin fixation. HistoChoice® produced comparable or improved staining for alkaline phosphatase. Acid phosphatase localization was better in formalin fixed specimens, but osteoclasts were visuralized more easily in HistoChoice® fixed specimens. As expected, immunohistochemical labeling was antibody dependent; some antibodies labeled better in HistoChoice® fixed specimens while others were better in formalin fixed specimens. Toluidine blue, Von Kossa, and alkaline phosphatase staining of sections fixed for 12 h produced sections that were similar to 4 h fixed sections. Fixation for 12 h preserved acid phosphatase activity better. Increasing fixation to 12 h affected immunolocalization differentially. Bone sialoprotein labeling in HistoChoice® fixed specimens was comparable to formalin fixed samples. On the other hand, after 12 h formalin fixation, osteocalcin labeling was comparable to HistoChoice®. For most histochemical applications, fixing murine bone specimens for 4 h with HistoChoice® yielded superior staining compared to formalin fixation. If immunohistochemical localization is desired, however, individual antibodies must be tested to determine which fixation process retains antigenicity better. In addition, there was no detectable difference in the intensity of fluorochrome labeling using either fixative. Finally, fixation duration did not alter the intensity of labeling.     

Involvement of subtypes γ and ε of protein kinase C in colon pain induced by formalin injection

Protein kinase C (PKC) has been widely reported to participate in somatic pain; however, its role in visceral pain remains largely unclear. Using a colon inflammatory pain model by intracolonic injection of formalin in rats, the present study was to examine the role of PKC in visceral pain and determine which subtypes may be involved. The colon pain behavior induced by formalin injection could be enhanced by intrathecal pretreatment with a PKC activator (PMA), and alleviated by a PKC inhibitor (H-7). Wide dynamic range (WDR) neurons in the L6-S1 spinal dorsal horn that were responsive to colorectal distension were recorded extracellularly. It was found that neuronal activity was greatly increased following formalin injection. Microdialysis of PMA near the recorded neuron in the spinal dorsal horn facilitated the enhanced responsive activity induced by formalin injection, while H-7 inhibited significantly the enhanced response induced by formalin injection. Western blot analysis revealed that membrane translocation of PKC-γ and PKC-ε, but not other subtypes, in the spinal cord was obviously increased following formalin injection. Therefore, our findings suggest that PKC is actively involved in the colon pain induced by intracolonic injection of formalin. PKC-γ and PKC-ε subtypes seem to significantly contribute to this process.     

Honey as a substitute for formalin?

Formalin has long been the standard fixative for clinical routines worldwide. After the Formaldehyde Standard became law in the US in 1987, as a result of increasing concerns about the potential carcinogenicity of formaldehyde, attempts have been made to find safer alternatives. Alcoholic formalin is a useful fixative, because in addition to fixation, dehydration also is begun. For centuries, honey has been known to be an antibacterial agent with the potential to preserve compounds without harmful effects on its users. We compared the effects of honey fixation with other routine fixatives using conventional histochemical and immunohistochemical staining methods. Our results demonstrated that tissues fixed in either honey or alcoholic formalin and 10% neutral buffered formalin (NBF) have similar histomorphology. Honey fixation showed minor histomorphological differences among the various tissues; however, it did not influence affect correct diagnostic conclusions. Our results suggested that honey can be used as a safe alternative to formalin in histopathology.     

Quantitative proteomic analysis of formalin–fixed,paraffin–embedded clear cell renal cell carcinoma tissue using stable isotopic dimethylation of primary amines

Background

Formalin-fixed, paraffin-embedded (FFPE) tissues represent the most abundant resource of archived human specimens in pathology. Such tissue specimens are emerging as a highly valuable resource for translational proteomic studies. In quantitative proteomic analysis, reductive di-methylation of primary amines using stable isotopic formaldehyde variants is increasingly used due to its robustness and cost-effectiveness.

Results

In the present study we show for the first time that isotopic amine dimethylation can be used in a straightforward manner for the quantitative proteomic analysis of FFPE specimens without interference from formalin employed in the FFPE process. Isotopic amine dimethylation of FFPE specimens showed equal labeling efficiency as for cryopreserved specimens. For both FFPE and cryopreserved specimens, differential labeling of identical samples yielded highly similar ratio distributions within the expected range for dimethyl labeling. In an initial application, we profiled proteome changes in clear cell renal cell carcinoma (ccRCC) FFPE tissue specimens compared to adjacent non–malignant renal tissue. Our findings highlight increased levels of glyocolytic enzymes, annexins as well as ribosomal and proteasomal proteins.

Conclusion

Our study establishes isotopic amine dimethylation as a versatile tool for quantitative proteomic analysis of FFPE specimens and underlines proteome alterations in ccRCC.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1768-x) contains supplementary material, which is available to authorized users.     

Modification of the Falck-Hillarp formaldehyde fluorescence method using the vibratome®: simple,rapid and sensitive localization of catecholamines in sections of unfixed or formalin fixed brain tissue

Summary Two modifications of the original Falck-Hillarp formaldehyde fluorescence technique are presented, both based on a recently introduced instrument, the Vibratome^®, which permits cutting of unembedded tissue with a section thickness down to 10 ,.The first modification involves sectioning of unfixed tissue at a temperature below +5°C, subsequent air drying and reaction with formaldehyde vapours. In the second procedure formalin fixed tissue is cut and processed as described above. It is essential that both formalin fixation and cutting of the fixed tissue takes place at a low temperature to avoid diffusion of the catecholamines.The results show that with both techniques central CA neurons can be visualized with a high degree of sensitivity. Furthermore, since the sections are free from fractures—a common problem in freeze-dried tissues—the Vibratome^® technique represents a valuable tool for mapping studies. It may also be added that since many steps of the original procedure are omitted the present techniques are also more rapid and simple. It is pointed out that using the Vibratome^® procedure on formalin fixed tissue, it will be possible to combine e.g. cholinesterase staining or Fink-Heimer silver impregnation or immunofluorescent studies with the Falck-Hillarp technique on serial or even on the same sections.     

慢痛模型——鼠注射formalin试验的最佳浓度

长期以来国内外研究疼痛与镇痛机理多应用的是急性痛的动物模型,但在临床上更多遇到的是持续性慢痛,因此要求制造持久的伤害性刺激的慢痛模型才接近临床实际。给动物注射formalin试验最早是在1977年,由Dubuisson和Dennis在鼠和猫身上进行研究的,浓度为5%,注射量50μl,注射后30～60min期间观察其反应,按出现时间先后有一个早反应(earlg response)和一个迟反应(late response),1984年     

DNIC-mediated analgesia produced by a supramaximal electrical or a high-dose formalin conditioning stimulus: roles of opioid and α2-adrenergic receptors

Background  

Diffuse noxious inhibitory controls (DNIC) can be produced by different types of conditioning stimuli, but the analgesic properties and underlying mechanisms remain unclear. The aim of this study was to differentiate the induction of DNIC analgesia between noxious electrical and inflammatory conditioning stimuli.     

Checklist of the smaller families of Opomyzoidea,Anthomyzidae, Asteiidae,Aulacigastridae, Clusiidae,Odiniidae, Opomyzidae and Periscelididae (Diptera)?of?Finland

A species checklist is presented for Finland covering seven smaller families of Opomyzoidea: Anthomyzidae, Asteiidae, Aulacigastridae, Clusiidae, Odiniidae, Opomyzidae and Periscelididae (Diptera).     

Trypanosomes of non-human primates from the National Centre of Primates, Ananindeua, State of Pará, brazil

Trypanosome infections were sought in 46 non-human primates captured principally in Amazonian Brazil. Twenty-two (47.8%) were infected with four Trypanosoma species: T. cruzi, T. minasense, T. devei and T. rangeli. These preliminary results confirmed the high prevalence and diversity of natural infections with trypanosomes in primates from Brazilian Amazon and were the first formal record of simian infections with trypanosomes in the State of Acre. The presence of T. cruzi-like and T. rangeli-like parasites are recorded in four new hosts.     

Compounds isolated at theDepartment of Biogenesis of Natural Substances,Institute of Microbiology,Czechoslovak Academy of Sciences,in 1954–1983

Substances studied at this department in 1954–1983 are reviewed; a total of 226 compounds are characterized in a tabular form. They include natural compounds as well as those prepared by biotransformation, by semisynthetic and synthetic methods.     

Abstracts of Papers,Conference of Young Scientists,Kol’tsov Institute of Developmental Biology,Russian Academy of Sciences,Moscow, Russia (December 21–22, 2009)

Reviews

Abstracts of Papers, Conference of Young Scientists, Kol’tsov Institute of Developmental Biology, Russian Academy of Sciences, Moscow, Russia (December 21–22, 2009)     

Compounds isolated at theDepartment of Biogenesis of Natural Substances,institute of microbiology,Czechoslovak academy of sciences,in 1984–1988

A total of 41 compounds studied at theDepartment of Biogenesis of Natural Substances between 1984 and 1988 are characterized and tabulated. They include natural, semisynthetic and synthetic compounds. 1st supplement to the paper Podojilet al. (1984).     

Review of the fauna of Rotatoria,Cladocera, and Copepoda of the basin of the Anadyr’ River

The zooplankton composition is studied in the thermokarst, glacial and meteorite lakes, channels, former riverbeds, and hollows in the basin of Anadyr’. We found 174 taxa: 78, Rotatoria, 55, Cladocera, and 41, Copepoda. The most diverse is the lake fauna: 51 taxa of Rotatoria, 48, Cladocera, and 37, Copepoda. The thermokarst Lake Maiorskoe hosts 68 taxa: 31, Rotatoria, 14, Cladocera, and 23, Copepoda, wheras the cold ultraoligotrophic Lake El’gygytgyn features only one species of Cyclop of the group scutifer Cyclops neymanae Strel., and Rotatoria and Cladocera are present as allochtonous forms. The Copepoda illustrate the relations of the Anadyr’ fauna with those of Europe, North America, and Japan.     

Distribution, crypticity, stability, and localization of α-L-fucosidase of mouse cauda epididymal sperm

Sperm‐associated and semen‐specific isoforms of α‐L ‐fucosidase are thought to function in fertilization in numerous organisms. Here, we report the localization, distribution, crypticity, and stability of this enzyme in mouse cauda epididymal sperm and cauda fluid. Western analysis revealed that the sperm‐associated α‐L ‐fucosidase is present as two isoforms (Mr ～49 and 56 kDa), whereas the cauda fluid α‐L ‐fucosidase shows a single band at 50 kDa. α‐L ‐Fucosidase activity was detected using the fluorogenic substrate 4‐MU‐FUC. Of the total α‐L ‐fucosidase activity recovered in the cauda epididymal contents, 74% was found in the cell‐free cauda fluid and about 7% was found in sperm cells. During capacitation or permeabilization, cryptic intracellular stores of soluble enzyme were released to the supernatant, while leaving bound enzyme concentrated within the small volume of sperm. Moreover, membrane‐associated enzyme activity was still detectable in acrosome‐reacted cells. Immunofluorescence studies support the presence of α‐L ‐fucosidase (originally localizing at the acrosomal area) at the equatorial segment after the acrosome reaction. α‐L ‐Fucosidase activity of both cauda fluid and sperm at 37°C, 5% CO₂ was relatively stable and detectable up to 72 hr. The stability and appearance of mouse sperm‐associated α‐L ‐fucosidase in the equatorial segment after the acrosome reaction suggest that α‐L ‐fucosidase may be involved in sperm–egg interaction. Mol. Reprod. Dev. 79: 208–217, 2012. © 2011 Wiley Periodicals, Inc.     

An inventory of the mires of Hokkaido,Japan—their development,classification, decline,and conservation

Hokkaido Island is located in the cool temperate zone, and its climate conditions facilitated the formation of a variety of wetland types, the majority of them peat-forming mires. Most of these remained in a natural state until the early 20th century. However, drainage and subsequent conversion mostly to agricultural land have since destroyed more than 70% of the original wetland ecosystems. This paper (1) provides an overview of mire types, (2) reviews the development process of mires in Hokkaido during the Holocene, (3) analyzes the causes of losses of wetland areas, and (4) gives a summary of the current conservation and management status. Basic mire types that have been described in other parts of the northern hemisphere can also be recognized in Hokkaido, although there are floristic differences, and the frequency and intensity of volcanic impact and tsunamis is higher than in most other regions with abundant mire formation. Mire formation started at various points during the postglacial period; a few mountain mires in southwest Hokkaido date back to the Lateglacial, but most mountain mires formed during the mid to late Holocene. Most lowland mires developed at altitudes below 20 m and were influenced by the Jomon transgression that peaked ca. 6,000 years BP. The largest lowland mires started forming after the sea retreated, and many are not older than ca. 3,000 years. In 1996, the total number of wetlands (including peat-forming mires, freshwater marshes, and saltmarshes) greater than 1 ha was 150, with a total area of 59,881 ha. In 1928, when many wetlands were yet undeveloped, their total area was 200,642 ha. Most losses occurred between the 1950s and 1970s, when post-war development economics promoted agriculture and large-scale reclamation projects. Currently, 90.7% of mountain wetlands are public land, while 81.3% of the lowland wetlands are private or in mixed ownership. The ownership condition affects the possibilities for the protection of complete mire complexes especially in the lowlands. For effective conservation of wetland ecosystems it is necessary to include the catchment area in the planning of protected areas.     

List of reviewers, 2017

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