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Centella asiatica (L.) leaf extract treatment during the growth spurt period enhances hippocampal CA3 neuronal dendritic arborization in rats

Mohandas Rao KG, Muddanna Rao S, Gurumadhva Rao S

Evid Based Complement Alternat Med, 2006 Sep;3(3):349-57.
PMID: 16951719

Centella asiatica (CeA) is a creeping plant growing in damp places in India and other Asian countries. The leaves of CeA are used for memory enhancement in the Ayurvedic system of medicine, an alternative system of medicine in India. In this study, we have investigated the effect during the rat growth spurt period of CeA fresh leaf extract treatment on the dendritic morphology of hippocampal CA3 neurons, one of the regions of the brain concerned with learning and memory. Neonatal rat pups (7 days old) were fed with 2, 4 or 6 ml kg(-1) body weight of fresh leaf extract of CeA for 2, 4 or 6 weeks. After the treatment period the rats were killed, their brains were removed and the hippocampal neurons were impregnated with silver nitrate (Golgi staining). Hippocampal CA3 neurons were traced using a camera lucida, and dendritic branching points (a measure of dendritic arborization) and intersections (a measure of dendritic length) were quantified. These data were compared with data for age-matched control rats. The results showed a significant increase in the dendritic length (intersections) and dendritic branching points along the length of both apical and basal dendrites in rats treated with 4 and 6 ml kg(-1) body weight per day of CeA for longer periods of time (i.e. 4 and 6 weeks). We conclude that the constituents/active principles present in CeA fresh leaf extract have a neuronal dendritic growth stimulating property; hence, the extract can be used for enhancing neuronal dendrites in stress and neurodegenerative and memory disorders.

Matched MeSH terms: Staining and Labeling
Fulltext Betulinic Acid inhibits growth of cultured vascular smooth muscle cells in vitro by inducing g(1) arrest and apoptosis

Vadivelu RK, Yeap SK, Ali AM, Hamid M, Alitheen NB

Evid Based Complement Alternat Med, 2012;2012:251362.
PMID: 23056140 DOI: 10.1155/2012/251362

Betulinic acid is a widely available plant-derived triterpene which is reported to possess selective cytotoxic activity against cancer cells of neuroectodermal origin and leukemia. However, the potential of betulinic acid as an antiproliferative and cytotoxic agent on vascular smooth muscle (VSMC) is still unclear. This study was carried out to demonstrate the antiproliferative and cytotoxic effect of betulinic acid on VSMCs using 3-[4,5-dimethylthizol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay, flow cytometry cell cycle assay, BrdU proliferation assay, acridine orange/propidium iodide staining, and comet assay. Result from MTT and BrdU assays indicated that betulinic acid was able to inhibit the growth and proliferation of VSMCs in a dose-dependent manner with IC(50) of 3.8 μg/mL significantly (P < 0.05). Nevertheless, betulinic acid exhibited G(1) cell cycle arrest in flow cytometry cell cycle profiling and low level of DNA damage against VSMC in acridine orange/propidium iodide and comet assay after 24 h of treatment. In conclusion, betulinic acid induced G(1) cell cycle arrest and dose-dependent DNA damage on VSMC.

Matched MeSH terms: Staining and Labeling
Fulltext Detection of Respiratory Viruses from ARTI Patients by xTAG RVP Fast v2 Assay and Conventional Methods

Kuan CS, Yew SM, Hooi PS, Lee LM, Ng KP

Malays J Med Sci, 2017 Oct;24(5):33-43.
PMID: 29386970 MyJurnal DOI: 10.21315/mjms2017.24.5.4

Introduction: Acute respiratory tract infections (ARTIs) are a major cause of morbidity and mortality in paediatric patients. Therefore, early detection of the viral aetiologies of ARTIs is essential for patient management and infection control. In this study, we evaluated the performance of a new multiplex polymerase chain reaction (PCR) assay (xTAG Respiratory Viral Panel [RVP] Fast v2) in the detection of respiratory viruses by comparing it with that of viral culture and direct immunofluorescence (IF) staining.
Methods: Nasopharyngeal swab and aspirate samples were collected prospectively from 199 patients who presented with ARTIs at the University Malaya Medical Centre (UMMC) in Kuala Lumpur, Malaysia during a 10-month period. The PCR assay was conducted in parallel with conventional culture and direct IF staining methods.
Results: The positive rate of the xTAG RVP Fast v2 assay (78.4%) in detecting respiratory viruses was higher than that of the viral isolation (7.5%) and direct IF (23.1%) methods. Using the xTAG RVP Fast v2 assay, human enterovirus/human rhinovirus (HEV/HRV) was the most frequently detected (46.2%). The xTAG RVP Fast v2 assay revealed mixed infection caused by two or three respiratory viruses in 40 specimens, and these were undetected by the viral isolation and direct IF methods.
Conclusion: The xTAG RVP Fast v2 assay was superior to conventional methods in the identification of common respiratory viruses, with higher sensitivity and shorter turnaround times for laboratory results.

Matched MeSH terms: Staining and Labeling
Fulltext Anti-Cancer Properties of Heterotrigona itama sp. Honey Via Induction of Apoptosis in Malignant Glioma Cells

Ahmad F, Seerangan P, Mustafa MZ, Osman ZF, Abdullah JM, Idris Z

Malays J Med Sci, 2019 Mar;26(2):30-39.
PMID: 31447606 MyJurnal DOI: 10.21315/mjms2019.26.2.4

Background: There has been increasing evidence showing that stingless bee honey exhibits anti-oxidant, anti-inflammatory and anti-cancer properties. Pharmacologically-active components in honey such as flavonoids and phenolic constituents are known to contribute to its medicinal benefits. To the best of our knowledge, this is the first study on evaluating anti-cancer effects of locally-produced Malaysian stingless bee honey from Heterotrigona itama sp. on malignant glioma cells.
Methods: Proliferation and apoptosis studies of U-87 MG cells following stingless bee honey treatment were carried out using MTS assay and acridine orange/propidium iodide dual staining, respectively.
Results: Results demonstrated time and dose-dependent cytotoxicity using 0.625%, 1.25% and 10% stingless bee honey (P < 0.05). IC50 values were calculated using cells treated with 10% stingless bee honey. It was also observed that 10% stingless bee honey induced nuclear shrinkage, chromatin condensation and nucleus fragmentation, indicating that cellular changes were consistent with the apoptotic characteristics of the cells.
Conclusion: These data provide a good basis for further evaluation of the medicinal properties of stingless bee honey from Heterotrigona itama sp. This source of honey may serve as a potential therapy for malignant glioma.

Matched MeSH terms: Staining and Labeling
Fulltext Comparison of dry eye parameters between diabetics and non-diabetics in district of Kuantan, Pahang

Aljarousha M, Badarudin NE, Che Azemin MZ

Malays J Med Sci, 2016 May;23(3):72-7.
PMID: 27418872 MyJurnal

INTRODUCTION: Diabetes may affect the human body's systems and organs, including the eye. Diabetic retinopathy is the 5th leading cause of blindness globally. Diabetic subjects demonstrated dry eye symptoms that were also supported by the low values of the clinical tests.
PURPOSE: This study aimed to compare the dry eye symptoms and signs between diabetics and non-diabetics and tear functions between diabetic subjects with and without dry eye.
METHODS: This retrospective study was based on the observation of 643 medical files. Using a convenience sampling method, 88 subjects were found to report diabetes mellitus. The information extracted from the files included: date of first examination, age at first visit, gender, past ocular history, systemic disease, symptoms of dry eye disease and details of clinical diagnostic signs. Non-contact lens wearers were excluded. A group of 88, age and gender matched, control subjects were included for this comparison study.
RESULTS: The percentage of dry eye symptoms was higher in diabetic subjects (15.9%) compared with non-diabetic subjects (13.6%; p<0.001). The percentage of dry eye symptoms was also higher in diabetics with dry eye (63%) than in diabetics without dry eye (36.9%; p<0.001). Tear break up time was significantly different between diabetics and non-diabetics (p<0.001) and between diabetics with and without dry eye (p=0.046). The corneal staining was significantly different between diabetic subjects with and without dry eye (p=0.028).
CONCLUSION: Dry eye symptoms were significantly associated with diabetics. Tear break up time was significantly shorter in diabetics with dry eye compared to diabetics without dry eye.
KEYWORDS: Diabetes mellitus; cornea; dry eye syndromes; signs and symptoms; tears
Study site: Klinik Kesihatan Jalan Hospital, Kuantan, Malaysia

Matched MeSH terms: Staining and Labeling
Modified fields' stain: ideal to differentiate Dientamoeba fragilis and Blastocystis sp

Ragavan AD, Govind SK

Parasitol Res, 2015 Mar;114(3):1163-6.
PMID: 25614298 DOI: 10.1007/s00436-014-4296-8

Dientamoeba fragilis, a trichomonad parasite is usually found in the gastrointestinal tract of human, and it is known to be the cause for gastrointestinal disease. The parasite is globally distributed and mostly found in rural and urban areas. The parasite is found in humans and nonhuman primates such as the macaques, baboons, and gorillas. Often, the parasite is confused with another largely found organism in stools called Blastocystis sp. especially when seen directly under light microscopy on culture samples containing both parasites. Both sometimes are seen with two nuclei with sizes tending to be similar which complicates identification. Stools were collected fresh from nine previously diagnosed persons infected with D. fragilis who also were found to be positive for Blastocystis sp. Samples were then cultured in Loeffler's medium and were stained with Giemsa, iron hematoxylin, and modified Fields' (MF) stain, respectively. D. fragilis was differentiated from Blastocystis sp. when stained with MF stain by the presence of a thinner outer membrane with clearly demarcated nuclei in the center of the cell whilst Blastocystis sp. had a darker and thicker stained outer membrane with the presence of two nuclei. The staining contrast was more evident with modified Fields' stain when compared with the other two. The simplicity in preparing the stain as well as the speed of the staining procedure make MF stain an ideal alternate. The modified Fields' stain is faster and easier to prepare when compared to the other two stains. MF stain provides a better contrast differentiating the two organisms and therefore provides a more reliable diagnostic method to precisely identify one from the other especially when cultures show mixed infections.

Matched MeSH terms: Staining and Labeling/methods*
Pseudocyst forms of Trichomonas vaginalis from cervical neoplasia

Afzan MY, Suresh K

Parasitol Res, 2012 Jul;111(1):371-81.
PMID: 22398830 DOI: 10.1007/s00436-012-2848-3

Trichomonas vaginalis, a flagellated protozoan parasite causes a variety of adverse health consequences in both men and women. The parasite exists in the trophozoite and the pseudocystic stage. The study reports for the first time that pseudocyst forms of T. vaginalis isolated from cervical neoplasia (CN) patients demonstrated distinct, different and significant in vitro growth profiles when grown in vitro cultures from day 1 up to day 5 (p<0.05, Mann-Whitney test) when compared with the same life cycle stages isolated from non-cervical neoplasia but symptomatic patients (NCN). Pseudocysts from CN and NCN isolates remained viable in distilled water until 3 h 10 min and 2 h 10 min, respectively. The nucleus of pseudocysts in CN isolates using acridine orange and DAPI showed more intense staining revealing higher nuclear content. The FITC-labeled Concanavalin A stained stronger green fluorescence with surface of pseudocysts in CN isolates showing more rough and creased surface with higher numbers of deep micropores with larger numbers of chromatin masses, vacuoles, and hydrogenosomes. The study confirms that pseudocystic stage from CN, despite the uniformity in appearance of being rounded and showing no motility without a true cyst wall under light microscopy, demonstrated different biochemical, surface, and ultrastructural properties. The study provides evidence that phenotypic variant forms of pseudocysts does exist and possibly does play a role in exacerbating cervical cancer.

Matched MeSH terms: Staining and Labeling/methods
Bone marrow and adipose stem cells can be tracked with PKH26 until post staining passage 6 in in vitro and in vivo

Ude CC, Shamsul BS, Ng MH, Chen HC, Norhamdan MY, Aminuddin BS, et al.

Tissue Cell, 2012 Jun;44(3):156-63.
PMID: 22402173 DOI: 10.1016/j.tice.2012.02.001

Tracking of transplanted cells has become an important procedure in cell therapy. We studied the in vitro dye retention, survival and in vivo tracking of stem cells with PKH26 dye. Sheep BMSCs and ADSCs were labeled with 2, 4 and 8 μmol of PKH26 and monitored for six passages. Labeled BMSCs and ADSCs acquired mean cumulative population doubling of 12.7±0.4 and 14.6±0.5; unlabeled samples had 13.8±0.5 and 15.4±0.6 respectively. Upon staining with 2, 4 and 8 μmol PKH26, BMSCs had retentions of 40.0±5.8, 60.0±2.9 and 95.0±2.9%, while ADSCs had 92.0±1.2, 95.0±1.2 and 98.0±1.2%. ADSCs retentions were significantly higher at 2 and 4 μmol. On dye retention comparison at 8 μmol and 4 μmol for BMSCs and ADSCs; ADSCs were significantly higher at passages 2 and 3. The viability of BMSCs reduced from 94.0±1.2% to 90.0±0.6% and ADSCs from 94.0±1.2% to 52.0±1.2% (p<0.05) after 24h. BMSCs had significant up regulation of the cartilage genes for both the labeled and the unlabeled samples compared to ADSCs (p<0.05). PKH26 fluorescence was detected on the resected portions of the regenerated neo-cartilage. The recommended concentration of PKH26 for ADSCs is 2 μmol and BMSCs is 8 μmol, and they can be tracked up to 49 days.

Matched MeSH terms: Staining and Labeling/methods*
Fulltext Detection of Entamoeba histolytica in experimentally induced amoebic liver abscess: comparison of three staining methods

Ning TZ, Kin WW, Mustafa S, Ahmed A, Noordin R, Cheong TG, et al.

Asian Pac J Trop Biomed, 2012 Jan;2(1):61-5.
PMID: 23569836 DOI: 10.1016/S2221-1691(11)60191-3

To compare the efficacy of three different tissue stains, namely haematoxylin and eosin (H&E), periodic-acid Schiff (PAS) and immunohistochemical (IHC) stains for detection of Entamoeba histolytica (E. histolytica) trophozoites in abscessed liver tissues of hamster.

Matched MeSH terms: Staining and Labeling/methods*
Prevalence of microsporidia in an indigenous Orang Asli community in Pahang, Malaysia

Lono A, Kumar GS, Chye TT

Trans R Soc Trop Med Hyg, 2010 Mar;104(3):214-8.
PMID: 19716577 DOI: 10.1016/j.trstmh.2009.07.006

Microsporidia are ubiquitous parasites thought to be closely related to fungi. Their presence in the environment means that humans are frequently exposed to infection. Stool samples were collected from 151 indigenous villagers from the eastern state of Pahang in 2005. The samples were concentrated with water-ether sedimentation, stained with modified trichrome stain and examined under oil-immersion microscopy. Thirty-two specimens (21.2%) were positive for microsporidia. Microsporidia were observed as ovoid or rounded ovoid shapes measuring approximately 1mum, with a bright pink outline containing a central or posterior vacuole. PCR amplification with specific primers on microscopy-positive specimens amplified Encephalitozoon intestinalis DNA from five of the ten specimens used.

Matched MeSH terms: Staining and Labeling/methods
Endometrial and endocervical secretion: the search for histochemical differentiation

Nieuwenhuizen L, Khalil MK, Venkatesh N, Othman NH

Anal. Quant. Cytol. Histol., 2006 Apr;28(2):87-96.
PMID: 16637511

To determine the ideal histochemical stain to differentiate between non-neoplastic and neoplastic endocervix and endometrium.

Matched MeSH terms: Staining and Labeling*
Evaluation of formalin-ether sedimentation and trichrome staining techniques: its effectiveness in detecting Entamoeba histolytica/dispar/moshkovskii in stool samples

Anuar TS, Al-Mekhlafi HM, Abdul Ghani MK, Abu Bakar E, Azreen SN, Salleh FM, et al.

J Microbiol Methods, 2013 Mar;92(3):344-8.
PMID: 23361047 DOI: 10.1016/j.mimet.2013.01.010

This study was conducted to evaluate two routinely microscopic diagnostic methods in comparison with single-round PCR assay as the reference technique to detect Entamoeba histolytica/dispar/moshkovskii. Examination was performed on 500 stool samples obtained from Orang Asli communities in different states of Malaysia using formalin-ether sedimentation, trichrome staining and single-round PCR techniques. Ninety-three stool samples were detected E. histolytica/dispar/moshkovskii positive by routine microscopy, while single-round PCR detected 106 positive samples. Additional positives detected by PCR assay were eventually confirmed to be negative by both microscopic techniques. Detection rate of E. histolytica/dispar/moshkovskii was highest in combination techniques (18.6%), followed by trichrome staining (13.4%) and formalin-ether sedimentation (11.2%) techniques. Single-round PCR detected 21.2% of the stool samples. The sensitivity and specificity of formalin-ether sedimentation and trichrome staining techniques compared to the reference technique were 31.1% (95% CI: 29.0-36.0) and 94.2% (95% CI: 89.8-98.9), and 53.8% (95% CI: 46.0-76.2) and 97.5% (95% CI: 92.8-99.1), respectively. However, the sensitivity [59.4% (95% CI: 48.9-78.5)] of the method increased when both techniques were performed together, but the specificity decreased to 92.4% (95% CI: 81.0-98.0). The agreement between the reference technique, trichrome staining and combination techniques were statistically significant by Kappa statistics (trichrome staining: K = 0.592, p < 0.05; combination techniques: K = 0.543, p < 0.05). Hence, the combination technique is recommended to be used as a screening method in the diagnosis of E. histolytica/dispar/moshkovskii infections either for clinical or epidemiological study.

Matched MeSH terms: Staining and Labeling/methods
Fulltext Organ-Specific Analysis of Morus alba Using a Gel-Free/Label-Free Proteomic Technique

Zhu W, Zhong Z, Liu S, Yang B, Komatsu S, Ge Z, et al.

Int J Mol Sci, 2019 Jan 16;20(2).
PMID: 30654535 DOI: 10.3390/ijms20020365

Morus alba is an important medicinal plant that is used to treat human diseases. The leaf, branch, and root of Morus can be applied as antidiabetic, antioxidant, and anti-inflammatory medicines, respectively. To explore the molecular mechanisms underlying the various pharmacological functions within different parts of Morus, organ-specific proteomics were performed. Protein profiles of the Morus leaf, branch, and root were determined using a gel-free/label-free proteomic technique. In the Morus leaf, branch, and root, a total of 492, 414, and 355 proteins were identified, respectively, including 84 common proteins. In leaf, the main function was related to protein degradation, photosynthesis, and redox ascorbate/glutathione metabolism. In branch, the main function was related to protein synthesis/degradation, stress, and redox ascorbate/glutathione metabolism. In root, the main function was related to protein synthesis/degradation, stress, and cell wall. Additionally, organ-specific metabolites and antioxidant activities were analyzed. These results revealed that flavonoids were highly accumulated in Morus root compared with the branch and leaf. Accordingly, two root-specific proteins named chalcone flavanone isomerase and flavonoid 3,5-hydroxylase were accumulated in the flavonoid pathway. Consistent with this finding, the content of the total flavonoids was higher in root compared to those detected in branch and leaf. These results suggest that the flavonoids in Morus root might be responsible for its biological activity and the root is the main part for flavonoid biosynthesis in Morus.

Matched MeSH terms: Staining and Labeling*
Fulltext A rapid and sensitive system for recovery of nucleic acids from Mycobacteria sp. on archived glass slides

A Talip B, Snelling WJ, Sleator RD, Lowery C, Dooley JSG

BMC Microbiol, 2018 11 26;18(1):196.
PMID: 30477427 DOI: 10.1186/s12866-018-1335-0

BACKGROUND: The field of diagnostics continues to advance rapidly with a variety of novel approaches, mainly dependent upon high technology platforms. Nonetheless much diagnosis, particularly in developing countries, still relies upon traditional methods such as microscopy. Biological material, particularly nucleic acids, on archived glass slides is a potential source of useful information both for diagnostic and epidemiological purposes. There are significant challenges faced when examining archived samples in order that an adequate amount of amplifiable DNA can be obtained. Herein, we describe a model system to detect low numbers of bacterial cells isolated from glass slides using (laser capture microscopy) LCM coupled with PCR amplification of a suitable target.
RESULTS: Mycobacterium smegmatis was used as a model organism to provide a proof of principle for a method to recover bacteria from a stained sample on a glass slide using a laser capture system. Ziehl-Neelsen (ZN) stained cells were excised and catapulted into tubes. Recovered cells were subjected to DNA extraction and pre-amplified with multiple displacement amplification (MDA). This system allowed a minimum of 30 catapulted cells to be detected following a nested real-time PCR assay, using rpoB specific primers. The combination of MDA and nested real-time PCR resulted in a 30-fold increase in sensitivity for the detection of low numbers of cells isolated using LCM.
CONCLUSIONS: This study highlights the potential of LCM coupled with MDA as a tool to improve the recovery of amplifiable nucleic acids from archived glass slides. The inclusion of the MDA step was essential to enable downstream amplification. This platform should be broadly applicable to a variety of diagnostic applications and we have used it as a proof of principle with a Mycobacterium sp. model system.

Matched MeSH terms: Staining and Labeling/instrumentation
Fulltext Colonial morphotypes and biofilm forming ability of Burkholderia pseudomallei

Koh SF, Tay ST, Puthucheary SD

Trop Biomed, 2013 Sep;30(3):428-33.
PMID: 24189672 MyJurnal

Burkholderia pseudomallei the causative agent of melioidosis, is being increasingly recognized as an important cause of morbidity and mortality in South East Asia. Biofilm formation of B. pseudomallei may be responsible for dormancy, latency and relapse of melioidosis. Based on the colonial morphology of the bacteria on B. pseudomallei selective agar medium, seven distinct morphotypes were identified. This study was conducted to assess the in vitro biofilm produced by B. pseudomallei and to investigate possible correlation between B. pseudomallei morphotypes with biofilm forming abilities of the isolates. Using a standard biofilm crystal violet staining assay, comparison was made between the biofilm forming ability of 76 isolates of B. pseudomallei and Burkholderia thailandensis ATCC 700388. Amongst the blood isolates, 30.2% were considered as high biofilm producers and 27.9% were low producers, 33.3% of the pus isolates were considered as high and 16% low biofilm producers. Most of the isolates were identified as morphotype group 1 which displayed a rough centre with irregular circumference on the agar medium. However, we did not find any correlation of B. pseudomallei morphotypes with biofilm forming abilities (p > 0.05). Additional studies are needed to identify internal and external factors which contribute to the high and low biofilm formation of B. pseudomallei.

Matched MeSH terms: Staining and Labeling/methods
Species identification of intestinal microsporidia using immunofluorescence antibody assays

Al-Mekhlafi MA, Fatmah MS, Anisah N, Azlin M, Al-Mekhlafi HM, Norhayati M

Southeast Asian J Trop Med Public Health, 2011 Jan;42(1):19-24.
PMID: 21323160

Abstract. The species identification of Enterocytozoon bieneusi and Encephalitozoon intestinalis is only possible using transmission electron microscopy (TEM), mo lecular techniques and immunofluorescence antibody assays (IFA). In this study, 50 positive and 50 negative fecal specimens for microsporidial spores using the Weber modified trichrome (WMT) staining technique were examined using IFA-MAbs. Of the 100 specimens examined, the microsporidial spores identified by IFA-MAbs were Enterocytozoon Bieneusi 42 (75%) Encephalitozoon intestinalis 7 (12.5%) and mixed infections 7 (12.5%). The sensitivity and specificity of IFA-MAbs in detecting microsporidial spores were 98% and 86%, respectively. The agreement between the WMT staining technique and IFA-MAbs was statistically significant by Kappa statistics (K = 0.840; p < 0.001). E. bieneusi was the commonest Microsporidia species isolated from the studied population; the presence of microsporidial spores detected by IFA-MAbs should be confirmed by other methods.

Matched MeSH terms: Staining and Labeling/methods
Comparative analysis between multilevel sectioning with conventional haematoxylin and eosin staining and immunohistochemistry for detecting nodal micrometastases with stage I and II colorectal cancers

Wong YP, Shah SA, Shaari N, Mohamad Esa MS, Sagap I, Isa NM

Asian Pac J Cancer Prev, 2014;15(4):1725-30.
PMID: 24641399

Management of patients with stage II colorectal carcinomas remains challenging as 20 - 30% of them will develop recurrence. It is postulated that these patients may harbour nodal micrometastases which are imperceptible by routine histopathological evaluation. The aims of our study were to evaluate (1) the feasibility of multilevel sectioning method utilizing haematoxylin and eosin stain and immunohistochemistry technique with cytokeratin AE1/AE3, in detecting micrometastases in histologically-negative lymph nodes, and (2) correlation between nodal micrometastases with clinicopathological parameters. Sixty two stage I and II cases with a total of 635 lymph nodes were reviewed. Five-level haematoxylin and eosin staining and one-level cytokeratin AE1/AE3 immunostaining were performed on all lymph nodes retrieved. The findings were correlated with clinicopathological parameters. Two (3.2%) lymph nodes in two patients (one in each) were found to harbour micrometastases detected by both methods. With cytokeratin AE1/AE3, we successfully identified four (6.5%) patients with isolated tumour cells, but none through the multilevel sectioning method. Nodal micrometastases detected by both multilevel sectioning and immunohistochemistry methods were not associated with larger tumour size, higher depth of invasion, poorer tumour grade, disease recurrence or distant metastasis. We conclude that there is no difference between the two methods in detecting nodal micrometastases. Therefore it is opined that multilevel sectioning is a feasible and yet inexpensive method that may be incorporated into routine practice to detect nodal micrometastases in centres with limited resources.

Matched MeSH terms: Staining and Labeling/methods*
Evaluation of gram-chromotrope kinyoun staining technique: its effectiveness in detecting microsporidial spores in fecal specimens

Salleh FM, Al-Mekhlafi AM, Nordin A, Yasin 'M, Al-Mekhlafi HM, Moktar N

Diagn Microbiol Infect Dis, 2011 Jan;69(1):82-5.
PMID: 21146718 DOI: 10.1016/j.diagmicrobio.2010.08.028

This study was conducted to evaluate the modification of the usual Gram-chromotrope staining technique developed in-house known as Gram-chromotrope Kinyoun (GCK) in comparison with the Weber Modified Trichrome (WMT) staining technique; as the reference technique. Two hundred and ninety fecal specimens received by the Microbiology Diagnostic Laboratory of Hospital Universiti Kebangsaan Malaysia were examined for the presence of microsporidial spores. The sensitivity and specificity of GCK compared to the reference technique were 98% and 98.3%, respectively. The positive and negative predictive values were 92.5% and 99.6%, respectively. The agreement between the reference technique and the GCK staining technique was statistically significant by Kappa statistics (K = 0.941, P < 0.001). It is concluded that the GCK staining technique has high sensitivity and specificity in the detection of microsporidial spores in fecal specimens. Hence, it is recommended to be used in the diagnosis of intestinal microsporidiosis.

Matched MeSH terms: Staining and Labeling/methods*
Modified Field's staining--a rapid stain for Trichomonas vaginalis

Afzan MY, Sivanandam S, Kumar GS

Diagn Microbiol Infect Dis, 2010 Oct;68(2):159-62.
PMID: 20846588 DOI: 10.1016/j.diagmicrobio.2010.06.005

Trichomonas vaginalis, a flagellate protozoan parasite commonly found in the human genitourinary tract, is transmitted primarily by sexual intercourse. Diagnosis is usually by in vitro culture method and staining with Giemsa stain. There are laboratories that use Gram stain as well. We compared the use of modified Field's (MF), Giemsa, and Gram stains on 2 axenic and xenic isolates of T. vaginalis, respectively. Three smears from every sediment of spun cultures of all 4 isolates were stained, respectively, with each of the stains. We showed that MF staining, apart from being a rapid stain (20 s), confers sharper staining contrast, which differentiates the nucleus and the cytoplasm of the organism when compared to Giemsa and Gram staining especially on parasites from spiked urine samples. The alternative staining procedure offers in a diagnostic setting a rapid stain that can easily visualize the parasite with sharp contrasting characteristics between organelles especially the nucleus and cytoplasm. Vacuoles are more clearly visible in parasites stained with MF than when stained with Giemsa.

Matched MeSH terms: Staining and Labeling/methods*
Fulltext Safety and Efficacy of Human Wharton's Jelly-Derived Mesenchymal Stem Cells Therapy for Retinal Degeneration

Leow SN, Luu CD, Hairul Nizam MH, Mok PL, Ruhaslizan R, Wong HS, et al.

PLoS One, 2015;10(6):e0128973.
PMID: 26107378 DOI: 10.1371/journal.pone.0128973

To investigate the safety and efficacy of subretinal injection of human Wharton's Jelly-derived mesenchymal stem cells (hWJ-MSCs) on retinal structure and function in Royal College of Surgeons (RCS) rats.

Matched MeSH terms: Staining and Labeling/methods

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