Sample Turnaround Times for Three Histoprocessing Methods

Sample Turnaround Times for Three Histoprocessing Methods Introduction Turnaround time for any pathological laboratory is very important which depends upon the preparation and diagnosis of the histopathologic material. The rapidity advantages the clinician to treat acutely ill patients and influence the work practice of the pathologist. With the advent of modernization tissue processing is modified from the point of tissue removal to embedding for instant histopathological diagnosis by various techniques or methods. After the surgical removal, the tissue undergoes preparatory protocol for preparation of sections which usually involves impregnation with a suitable supporting medium. The stages of tissue processing include fixation, dehydration, clearing, impregnation and embedding for designated durations of time to ensure completion of the procedure. Culling 1974, Bancroft Gamble 2002 The reproducibility and relatively low expense attached to the most commonly employed method continues to recommend it as a valuable tool after nearly 100 years of existence. But with the demand of faster or early reporting, newer techniques like rapid manual and microwave processing are getting introduced. Each of them is unique with their own set of advantages and disadvantages. The conventional tissue processing is reliable and cost effective but time consumption, reagent toxicity and delay in providing diagnosis are the major disadvantages. The rapid manual tissue processing has major disadvantages like the use of noxious chemicals, greater degree of tissue distortion and shrinkage which led to exploration for new short processing schedules. The microwave tissue processing eliminates the use of noxious chemicals, causes lesser distortion of tissue and has shorter processing time but the cost involved in instrumentation is very high. (Panja et al. 2007) Microwaves were invented by Percy Spencer in 1945 which work on the principle of producing heat by oscillating or exciting polar molecules. The microwave irradiation forces dipolar molecules of proteins to rotate through 180Â ° at the rate of 2.45 billion cycles per second (Srinivasan, Sedmak Jewell 2002, Bancroft Gamble 2002). These excited molecules due to kinetics cause collision with adjacent molecules resulting in transfer of rotational energy. This friction causes production of heat within the material itself leading to accelerated diffusion of processing fluids hence faster processing is possible. The advantages associated with microwave processing led to the production of commercially available microwaves specifically designed for tissue processing, however, the cost involved in these is very high (Leong 2004, Rohr et al. 2001). Domestic microwaves are readily available, affordable and had been used for tissue processing with good results earlier by some authors. Thus, the aim of the present study was to compare and analyse the efficacy of three histoprocessing methods and to determine the impact on turnaround times of tissue processing by these three methods. Materials and method In the present study, 60 specimens were selected randomly. The soft tissue specimen fixed in 10% NBF for 24h were included in the study and hard tissues like cartilage, bone and tooth were not included in the study. The gross features of the specimen were recorded and tissues were cut into three pieces of approximately same size to be processed by three methods. The sections obtained after processing were subsequently stained with HE by routine and microwave staining method. The stained slides in each group processed by three techniques were randomly numbered for a blind study and circulated among four observers referred as O1-O4. The observers graded each parameter on the format given in appendix D by following specific criteria as given in appendix C. Methodology Microwave tissue processing The microwave oven was calibrated as the microwave energy is non uniform within the chamber. Thus, hot and cold spots were detected in the chamber with the use of thermal paper sheet instead of the use of extra water load as suggested by various authors. The cold spot provided the most consistent results every time. The technique was self-standardized by trial and error method in the LG domestic Microwave (Model no. MS-285SD). The microwave was operated at the maximum output power of 40% (approximately 360 W) with rotating tray and ring removed. The cut piece from a fixed tissue sample was placed in a plastic tissue cassette and water washed in running tap water so that tissue was free of formalin. The tissue was irradiated in 200ml of 100% methanol and 200ml of 100% isopropyl alcohol for dehydration at cold spot for 2 cycles of 10 minutes each respectively in the microwave. After dehydration tissue was impregnated in 200ml of molten paraffin wax for 2 cycles at cold spot of 10 min each and was embedded in paraffin wax. Conventional Tissue Processing The cut piece from a fixed tissue sample was placed in a metal tissue cassette and water washed in running tap water so that tissue should be free of formalin. The tissue was dehydrated in 70% alcohol (one change), 90% alcohol (one change) and 100% alcohol three changes of 1 h each respectively. After dehydration tissue was cleared in two changes of xylene of 1 h each. Finally, tissue was impregnated in 2 changes of molten paraffin wax for 1 h each and was embedded. Rapid Manual Tissue Processing The cut piece from a fixed tissue sample was water washed in running tap water so that tissue should be free of formalin, after that tissue was wrapped in filter paper and dehydrated in 95% alcohol, 100% alcohol for 20 min on a stir plate. The dehydrated tissue was cleared in xylene for 20 min on a stir plate 20 min on a stir plate. Tissue was impregnated in 2 changes of molten paraffin wax of 1h each and was then embedded. The microwave processed tissue were stained as given in table no. ____. Conventional and rapid manual processed tissues were stained as given in table no.___________. Statistical Analysis The values obtained from different observers after assessment of sections processed by the three techniques were subjected to statistical analysis by Kruskal Wallis Test. One way ANOVA (Analysis of variance) was used for comparing mean shrinkage in tissues processed by the three histoprocessing methods. The P value Results All observers were assumed to be reliable as the Cronbach’s reliability test was statistically significant. Complete concordance was found amongst all pathologists in most of the cases. Hence, observer 1 was randomly selected for further analysis. The histopathological evaluation of the epithelium, fibrous tissues and glandular tissue revealed that the nuclear cytoplasmic contrast was good and cellular outline was distinct in tissues processed by microwave assisted technique followed by conventional processing and rapid manual processing techniques. The stroma was good with distinct cellular outline. The secretory products can be easily appreciated and the RBCs, inflammatory cells were intact. The results were statistically non-significant as observed by Kruskal Wallis test. The colour intensity of the tissues graded by four observers revealed that the microwave sections were crisper and there was a good contrast between the hematoxyphilic and eosinophillic areas. Though some slides were not visualised up to the mark, all the three histoprocessing methods were comparable to each other. One way Analysis of Variance (ANOVA) revealed a significant value on comparison of the tissue shrinkage processed by the three techniques. The dimensions of the tissues were recorded before dehydration and paraffin embedding. The mean percentage of shrinkage in rapid manual technique was significantly higher as compared to the other two techniques whereas statistically non-significant value was obtained on comparing conventional and microwave method of tissue processing. Discussion Microwaves are electromagnetic waves(Microwave Processing Techniques for Microscopy) which causes oscillation and excitation of polar molecules which are usually dipolar molecules of proteins in tissues. The excited molecules cause collision with adjacent molecules due to kinetics producing friction and causes production of heat within the material itself. The heat produced enhances the rate of diffusion of fluids to permeate into the tissues. The rise in temperature decreases the viscosity of processing fluids that facilitates diffusion. Therefore it is theoretically possible to fasten the tissue fixation and processing. This has resulted in a substantial reduction in the basic steps of histoprocessing, thereby reducing turnaround time and providing same day diagnosis. The applications of microwaves are extensive which includes tissue fixation, stabilization of large specimens, tissue processing for light and electron microscopy, histochemical and immunohistochemical staining. Microwave tissue processing technique was introduced by Boon and Kok in 1985 (Leong 2004) but the potential application of microwave energy was first recognized by Mayers in 1970 who successfully fixed tissue with a microwave generator (Kok, Visser Boon 1988). Boon et al. (1986) reported that it was possible to produce significant acceleration of tissue processing by using microwave radiation. Visinoni et al. (1998) first described the tissue processor that completed the processing in 30-120 min, thus reducing the processing time from 24 h to just 1-2 h providing early reporting and easy patient management. Thus, the aim of the present study was to compare the cytoplasmic and nuclear details as well as staining characteristics of tissue sections processed by conventional, rapid manual and microwave techniques. The noxious chemicals used in conventional tissue processing were replaced in microwave tissue processing. In the microwave processing in contrast to conventional tissue processing, isopropyl alcohol was replaced by methanol as dehydrating agent and xylene by isopropyl alcohol as intermediate agent. Molten paraffin wax remained the impregnating and embedding medium for both the techniques. The reagent selection was in consonance with Babu, Malathi and Mangesh (2011) who also used methanol, isopropyl alcohol and molten paraffin wax for microwave tissue processing. Microwave radiation produced when enter the chamber it is reflected by the chamber walls until these gets absorbed by the material placed inside the chamber (Wong 2011). However, the spreading is not even throughout the chamber leading to formation of hot and cold spot zones. (Kok, Boon Smid 1993, Thostenson Chow 1999, Rutgers 2013). Hence, hot cold spots should be detected to achieve consistent results. Various authors have described methods for detection of hot and cold spots. Microwave processing was self-standardized by trial and error method in which the hot and cold spots were detected by using a damp thermal paper Kok, Boon and Smid (1993). All the procedures in the microwave were carried out in the cold spot zones as suggested by Sharp and Paperiello (1971), Benard (1974), Rangell and Keller (2000) in their respective studies. Microwavable plastic tissue cassettes were used for microwave tissue processing which are cheap and reusable as metallic utensils are contraindicated in the microwave because the electric fields of the waves produced by microwave magnetron are completely reflected at the same frequency by metals which can lead to sparking. (Vollmer 2004). In the present study, the staining protocol for microwave was followed as given by Babu, Malathi and Mangesh (2011) which included the stains used to be accelerated in the microwave. Kayser and Bubenzer (1990) used domestic microwave oven for acceleration of the various stains which also included HE stain. Valle (1986), Moorlag, Boon and Kok (1987) and Mathai et al. (2008) modified various special stain protocols for microwave and concluded that microwave did not produce any deleterious effects on staining. In our study, the three pieces of tissue processed by three techniques sectioned by a soft tissue microtome and stained as per their respective protocols were evaluated. We adopted the criterion for evaluation of tissue sections given by Kango and Deshmukh (2011). The overall quality of the tissue sections processed by microwave and manual methods was comparable. The microwave processed sections had same or similar cytoplasmic and nuclear details with good erythrocyte integrity and lymphocytic appearance than the manual methods. Similar results are given by Mathai et al. (2008), Morales et al. (2002), Bhuvanamha et al. (2013), Panja et al. (2007), Boon et al. (1986), Kok et al. (1988). We also observed that the stroma in some cases was slightly more condensed focally in microwave processed tissue sections which is similar to the findings reported by Boon, Kok and Ouwerkerk-Noordam (1986) This lead to the erroneous categorization of these cases as indistinct in studies by Kango and Deshmukh (2011). Since our criterion was adopted from the above mentioned study we also placed focal condensation of stroma as indistinct. In contrast Kok, Visser and Boon (1988) refuted the importance of focal condensation of stroma in diagnostic pathology. The colour intensity of the tissues graded by four observers revealed that the microwave sections were crisper and there was a good contrast between the hematoxyphilic and eosinophillic areas. The microwave processed tissues showed an increased reaction to HE. The sections stained were slightly more eosinophilic as compared to the manual techniques. Similar findings are reported by Hopwood et al. (1984), Boon et al. (1986), Chaudhari, Chattopadhyay and Dutta (2000), Leong and Price (2004), Panja et al. (2007), Mathai et al. (2008), Babu, Malathi and Mangesh (2011). Hopwood et al. (1984) suggested that this eosinophilia could be easily corrected by altering the stain composition or staining time in eosin. In contrast Leong Price (2004) observed that eosinophilia of the cytoplasm was advantageous as it produced good nuclear cytoplasmic contrast and enhancement of the cellular features. The dysplastic features i.e. hyperchromatism, pleomorphism of tumor cells and mitotic figures were easily appreciable in the microwave processed tissue sections of malignancy. There was also an easy appreciation of the giant cells in the tissues of Central Giant cell Granuloma and tubercular lymphadenitis processed by microwave processing technique. Rapid processing of histopathologic material is becoming increasingly desirable for intraoperative consultations and timely diagnosis. We found positive impact on turnaround time in microwave method as the time taken for block preparation from fixed tissue was 1h as compared to conventional method (9h) and rapid manual method (3h). In some cases, proper diagnosis could not be reached as the size of the tissue was small and the sample was not representative of the site. Similar difficulties were also encountered by Suri et al. (2006), Kango and Deshmukh (2011) in their respective studies. As assessed in our study, the effects of the three methods of histoprocessing on cytoplasmic and nuclear details of epithelial, fibrous and glandular tissue showed no statistically significant variation. The microwave technique was comparable or slightly better than the manual methods. Conclusion The applications and versatility of microwave processing methods are unattainable with conventional procedures. The method reported herein reproducibly yields similar histologic quality to that provided by conventional processing. It has many advantages including feasibility, safety and elimination of noxious chemicals that might be used for improvement in the practice of the histopathology laboratory, permitting the preparation of diagnostic material within a day. Domestic microwaves are easily available and cost effective but have certain notable disadvantages like uneven heating and inability to record and maintain temperature within the chamber. Further exploration in the field is required for development of cost effective microwave histoprocessors for histopathology which provide similar histologic material for rapid diagnosis and reporting. References

John F. Kennedy :: essays research papers fc

John F. Kennedy John Fitzgerald Kennedy was born on May 29, 1917 in the Boston suburb of Brookline. Kennedy was the son of Joseph P. Kennedy a formerambassador to Great Britain. Kennedy was much like his father, possesing a delightful sense of humor, a strong family loyalty, a concern for the state of the nation, endless vitality and a constant air of confidence no matter how dire the situation (Kennedy, Sorensen, Harper & Row, New York 1965, Page 18). Growing up in a priviliged household and graduating with honors from Harvard. He served as an assistant to his father (1938), naval officer (1941-1945), journalist (1941 and 1945) and Congressman (1947-1953), he had traveled to every major continent and talked with the presidents and prime ministers, of some thirty-seven countries. In 1952 he was elected to the United States Senate and in 1953 he married Jaqueline Bouvier. However one year later a spinal operation brought him to the edge of death's door, causing him to deeply reflect on his character (Sorensen 28). After his dangerous operation he researched and wrote a book, about democracy. The next year narrowly missing the Vice Presidential nomination of his party, Kennedy emerged as a national figure in large demand. "John Kennedy was not one of the Senate's great leaders" (Sorensen 43). Very few laws of great importance bear his name. Even after his initial â€Å" traditionally' inactive freshman year in the Senate, his chances for major contributions to the Senate excluding his stances on fair labor reform and against rackets, were constantly diminished of his Presidential campaign. His voting record reflects his open minded views, and strengthed beliefs. He was well liked and respected by many Senators. Kennedy was regarded for his eagerness and cool logic in debate situations His only real â€Å"enemy† was Senator Joseph McCarthy of Wisconsin (Sorensen 45).McCarthy's rough and wide-ranging hunts for Red, "pinks" and headlines had stomped on the freedoms of people who had not committed a crime, and Kennedy was too rational and reasonable a man to remain indiferent to the extremism known as Mcarthyism. Kennedy often was a thorn in McCarthy's side obstructing many of McCarthy's personal choices for various offices and by serving on certain committies of which McCarthy was chairman, such as the Government Operations Committee (Sorensen 46). Kennedy's political philosophy revoloved around the idea that one could not allow the pressures of party responisbility to cloud ones personal responsibility. Meaning after all was said and done that the decision falls upon yourself to make the choice regardless of what your party platform was. Of course the platfrom had significant merit, nevertheless it still came down to

Logic Gates

A Project Report On WORKING OF LOGIC GATES SUBMITTED BY:- Shamil Choudhury ROLL__________ No_____________ Karimganj Junior College Of Science CONTENTS†¦Ã¢â‚¬ ¦.. TOPICS Page No 1. CERTIFICATE1 2. INTRODUCTION 2 3. THEORY3-4 4. EXPERIMENT5-6 5. RESULTS 7 6. CONCLUSION. 7 7. ACKNOWLEDGEMENT 8 8. REFERENCE 9 INTRODUCTION A gate is a digital circuit that follows certain logical relationship between the input and output voltages. Therefore, they are generally known as logic gates — gates because they control the flow of information. The five common logic gates used are NOT, AND, OR, NAND, NOR. Each logic gate is indicated by a symbol and its function is defined by a truth table that shows all the possible input logic level combinations with their respective output logic levels. Truth tables help understand the behavior of logic gates. These logic gates can be realized using semiconductor devices. Here we will investigate on the working of first three logic gates i. e. OR, AND, and NOT gate. OBJECTIVE OF THE PROJECT To demonstrate the working of logic gates using torch bulb, battery and switches. APPARATUS REQUIRED To demonstrate the working of the logic gates we require the following apparatus:- i) A BATTERY OF 6 V. ii) TWO SWITCHES (KEYS). iii) TORCH BULB. iv) CONNECTING WIRES THEORY There are three basic logic gates: 1. The OR Gate * An OR gate has two or more inputs with one output. It operates in such a way that a output is high (i. e. 1) if one of the two inputs (or both the inputs) is high. The symbol of OR Gate is It works on the following truth table:- 2. The AND GATE * An AND gate has two or more inputs and one output. It operates in such a way that the output is high (i. . 1) when both the inputs are high. The symbol of AND Gate is It works on the following truth table:- 3. The NOT Gate * The NOT gate has single input and single output. The output is not the same as input. If the input is low (i. e. 0), the output is high (i. e. 1). On the other hand, the output will be low (i. e. 0) if the input is high (i. e. 1). In other words it performs a negation operation on the input and call ed the invertor. The symbol of NOT Gate is It works on the following truth table:- PROCEDURE (1) Working of OR Gate The electric circuit diagram is as shown Using the above circuit diagram we proceed as follows: (i) The switches S1 and S2 are not pluged and the glow of the bulb is recorded (do not glow) . (ii) Now the switch S1 is plugged (not S2), it is found that the bulb glows. (iii) Now the switch S2 is plugged (not S1), it is observed that the bulb glows. (iv) Both the switches are plugged, the bulb glows. (2) Working of AND gate The electric circuit diagram is shown Using the above circuit diagram we proceed as follows: (i) The switches S1 and S2 are not plugged and the glow of the bulb is recorded (do not glow) . ii) Now the switch S1 is plugged (not S2), it is found that the bulb does not glow. (iii) Now the switch S2 is plugged (not S1), it is observed that the bulb does not glow. (iv) Both the switches are plugged, the bulb glows. 3. Working of NOT gate The electric circuit diagram is shown Using the above circuit diagram we proceed as follows: (i) The switch S is not plugged and the glow of the bulb is recorded(the bulb glows) . (ii) The switch S is plugged and the glow of the bulb is recorded(the Bulb does not glow). RESULT 1. Results for the OR gate S1 | S2| Bulb glows| Off On Off On| Off Off On On| No Yes Yes Yes| 2. Results for the AND gate S1 | S2| Bulb glows| Off On Off On| Off Off On On| No No No Yes| 3. Results for the NOT gate S| Bulb glows | Off On| Yes No| CONCLUSION Thus we find that the logic gates works according to their corresponding truth table. ACKNOWLEDGEMENT I extend my sincere thanks to our Dr Sujit Tiwari for his advice and help in the matter of preparing the project. I also express my deep sense of gratitude to my friends forextending all possible help in supplying project information, which I have used while preparing the project. They have given lots of information for preparation of the report I sought. My parents have supplied me all kinds of necessary materials and information for the report files. Hence, I am grateful to them also. REFERENCE [1] Concept of Physics – H. C. Verma. [2] Comprehensive Practical Physics.

A Biography of the Venerable Bede

The Venerable Bede was a British monk whose works in theology, history, chronology, poetry, and biography have led him to be accepted at the greatest scholar of the early medieval era. Born in March of 672 and having died on May 25, 735 in Jarrow, Northumbria, UK, Bede is most famous for producing the Historia ecclesiastica (Ecclesiastical History), a source essential for our understanding of the Anglo-Saxons and the Christianisation of Britain in the era before William the Conqueror and the Norman Conquest, earning him the title of the Father of English history. Childhood Little is known of Bedes childhood, other than he was born in March of 672 to parents living on land belonging to the newly founded Monastery of St. Peter, based in Wearmouth, to which Bede was given by relatives for a monastic education when he was seven. Initially, in the care of Abbot Benedict, Bedes teaching was taken over by Ceolfrith, with whom Bede moved to the monasterys new twin-house at Jarrow in 681. The Life of Ceolfrith suggests that here only the young Bede and Ceolfrith survived a plague which devastated the settlement. However, in the aftermath of the plague the new house regrew and continued. Both houses were in the kingdom of Northumbria. Adult Life Bede spent the rest of his life as a monk at Jarrow, first being taught and then teaching to the daily rhythms of monastic rule: for Bede, a mixture of prayer and study. He was ordained as a Deacon aged 19 – at a time when Deacons were supposed to be 25 or over – and a priest aged 30. Indeed, historians believe Bede left Jarrow only twice in his relatively long life, to visit Lindisfarne and York. While his letters contain hints of other visits, there isnt any real evidence, and he certainly never traveled far. Works Monasteries were nodes of scholarship in early medieval Europe, and there is nothing surprising in the fact that Bede, an intelligent, pious and educated man, used his learning, life of study and house library to produce a large body of writing. What was unusual was the sheer breadth, depth, and quality of the fifty plus works he produced, covering scientific and chronological matters, history and biography and, perhaps as expected, scriptural commentary. As befitted the greatest scholar of his era, Bede had the chance to become Prior of Jarrow, and perhaps more, but turned the jobs down as they would interfere with his study. The Theologian: Bedes biblical commentaries – in which he interpreted the bible mainly as an allegory, applied criticism and tried to solve discrepancies – were extremely popular in the early medieval period, being copied and spread – along with Bedes reputation – widely across the monasteries of Europe. This dissemination was helped by the school of Archbishop Egbert of York, one of Bedes pupils, and later by a student of this school, Alcuin, who became head of Charlemagnes palace school and played a key role in the Carolingian Renaissance. Bede took the Latin and Greek of the early church manuscripts and turned them into something the secular elites of the Anglo-Saxon world could deal with, helping them accept the faith and spread the church. The Chronologist Bedes two chronological works - De temporibus (On Times) and De temporum ratione (On the Reckoning of Time) were concerned with establishing the dates of Easter. Along with his histories, these still affect our style of dating: when equating the number of the year with the year of Jesus Christs life, Bede invented the use of A.D., The Year Of Our Lord. In stark contrast to dark age cliches, Bede also knew the world was round, the moon affected tides and appreciated observational science. The Historian In 731/2 Bede completed the Historia ecclesiastica gentis Anglorum, the Ecclesiastical History of the English People. An account of Britain between the landings of Julius Caesar in 55/54 BC and St. Augustine in 597 AD, its the key source on the Christianisation of Britain, a mixture of sophisticated historiography and religious messages containing details simply not found elsewhere. As such, it now overshadows his other historical, indeed all his other, works and is one of the key documents in the entire field of British history. Its also lovely to read. Death and Reputation Bede died in 735 and was buried at Jarrow before being re-interred inside Durham Cathedral (at the time of this writing the Bedes World museum in Jarrow have a cast of his cranium on display.) He was already renowned among his peers, being described by a Bishop Boniface as having shone forth as a lantern in the world by his scriptural commentary, but is now regarded as the greatest and most multi-talented scholar of the early medieval era, perhaps of the entire medieval era. Bede was sainted in 1899, thus giving him the posthumous title of Saint Bede the Venerable. Bede was declared venerable by the church in 836, and the word is given on his tomb in Durham Cathedral: Hic sunt in fossa bedae venerabilis ossa (Here are buried the bones of the Venerable Bede.) Bede on Bede The Historia ecclesiastica finishes with a short account of Bede about himself and a list of his many works (and is actually the key source about his life that we, much later historians, have to work with): Thus much of the Ecclesiastical History of Britain, and more especially of the English nation, as far as I could learn either from the writings of the ancients, or the tradition of our ancestors, or of my own knowledge, has, with the help of God, been digested by me, Bede, the servant of God, and priest of the monastery of the blessed apostles, Peter and Paul, which is at Wearmouth and Jarrow; who being born in the territory of that same monastery, was given, at seven years of age, to be educated by the most reverend Abbot Benedict, and afterwards by Ceolfrid; and spending all the remaining time of my life in that monastery, I wholly applied myself to the study of Scripture, and amidst the observance of regular discipline, and the daily care of singing in the church, I always took delight in learning, teaching, and writing. In the nineteenth year of my age, I received deacons orders; in the thirtieth, those of the priesthood, both of them by the ministry of the most reverend Bishop J ohn, and by the order of the Abbot Ceolfrid. From which time, till the fifty-ninth year of my age, I have made it my business, for the use of me and mine, to compile out of the works of the venerable Fathers, and to interpret and explain according to their meaning... Source Bede, Ecclesiastical History of the English People. Penguin Classics, D. H. Farmer  (Editor, Introduction), Ronald Latham (Editor), et al., Paperback, Revised edition, Penguin Classics, May 1, 1991.