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استفسار عن ال electrophoresis??

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  • استفسار عن ال electrophoresis??

    السلام عليكم ورحمة الله وبركاته
    منتدى العزيز انا دورت على موضوع elerctrophoresis من يعرف اي شى يكتبه لى ولكم جزيل الشكر
    إلهي كيف أدعوك وأنا أنا ، وكيف أقطع رجائي منك وأنت أنت ، إلهي اذالم أسألك فتعطيني فمن ذا الذي أسئله فيعطيني ، إلهي إذا لم أدعك فتستجيبَ لي فمن ذا الذي أدعوه فيستجيبُ لي ، إلهي إذا لم أتضرع اليك فترحمني فمن ذا الذي أتضرعُ إليه فيرحمني ، إلهي فكما فلقت البحر لموسى عليه السلام ونجيته ، أسـألك أن تـصلـي عـلى محمدٍ وآلـه وأن تنجيـنـي مما أنا فـيـه وتفـرّجَ عـني فرجاً عـاجلاً غير آجـلٍ بفضلكَ ورحمتكَ يا أرحم الراحمين

  • #2
    السلام عليكم
    شو مالكم يا اعضائنا كلكم ما حد عندو ولا معلومة معلش انا مغلباكم بس شدو حالكم شوي
    بدي اي معلومة Enzyme electrophrosis
    إلهي كيف أدعوك وأنا أنا ، وكيف أقطع رجائي منك وأنت أنت ، إلهي اذالم أسألك فتعطيني فمن ذا الذي أسئله فيعطيني ، إلهي إذا لم أدعك فتستجيبَ لي فمن ذا الذي أدعوه فيستجيبُ لي ، إلهي إذا لم أتضرع اليك فترحمني فمن ذا الذي أتضرعُ إليه فيرحمني ، إلهي فكما فلقت البحر لموسى عليه السلام ونجيته ، أسـألك أن تـصلـي عـلى محمدٍ وآلـه وأن تنجيـنـي مما أنا فـيـه وتفـرّجَ عـني فرجاً عـاجلاً غير آجـلٍ بفضلكَ ورحمتكَ يا أرحم الراحمين

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    • #3
      http://www.science-projects.com/EnzElect.htm يا اختى افتحى الرابط ده

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      • #4
        للعضو hard_times_24 شكرا على الرابط بس اذا فيه غيره بكون احسن. وشكرا مره ثانييييييييييييييه
        \********s and Settings\Administrator

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        • #5
          شكرا لك يا اخى العزيز وبارك الله فيك
          إلهي كيف أدعوك وأنا أنا ، وكيف أقطع رجائي منك وأنت أنت ، إلهي اذالم أسألك فتعطيني فمن ذا الذي أسئله فيعطيني ، إلهي إذا لم أدعك فتستجيبَ لي فمن ذا الذي أدعوه فيستجيبُ لي ، إلهي إذا لم أتضرع اليك فترحمني فمن ذا الذي أتضرعُ إليه فيرحمني ، إلهي فكما فلقت البحر لموسى عليه السلام ونجيته ، أسـألك أن تـصلـي عـلى محمدٍ وآلـه وأن تنجيـنـي مما أنا فـيـه وتفـرّجَ عـني فرجاً عـاجلاً غير آجـلٍ بفضلكَ ورحمتكَ يا أرحم الراحمين

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          • #6
            اهلا بك اختي الكريمة ..موضوعك يتعلق بالكميستري ..سوف يتم نقل موضوعك وشكرآ
            http://www.arabslab.com/vb/uploaded/316_11191621547.jpg

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            • #7
              Electrophoresis of Enzymes

              Electrophoresis of Enzymes



              --------------------------------------------------------------------------------

              INTRODUCTION. The electrophoresis of active enzymes is a powerful tool because it tells you a lot about the status of the genes in the chromosomes of an organism, because enzymes, like all proteins, are the products of genes. There are several types of mutations in DNA, but the only ones that count are those that affect enzymatic activity.

              For example there are silent mutations which are base changes that do not affect the primary structures of proteins. Most often these mutations are in the third position of the genetic reading frame (and less frequently in the first position). Read about the "wobble hypothesis" for an historical perspective on this.
              However if a particular base has been substituted by another (a point mutation), AND the codon specifies an amino acid different from the previous, then we have a mutation which may or may not affect the enzyme' activity.
              It will not affect the activity if the location is in a portion of the polypeptide that is neither involved in the active site nor in the manner in which the polypeptide is folded. Often, for example a hydrophobic amino acid can be exchanged for another one because both have many of the same properties and are often found at the center of the finished protein - away from the surrounding water and helping to display the remainder of the enzyme and its active site to the water. These variants are called iso-enzymes (or isozymes, for short). However, the molecular weight of the enzyme will be slightly different, and if the substitution causes a slight change in charge to the molecule as a whole, then the molecule will move a bit differently in an electric field (electrophoresis). These isozymes are called "electromorphs."
              Activity will be affected especially if the amino is charged when the original was not or was of the opposite charge. Usually this mutation perturbs the working of the active site or to the imagined assembly line substratum. Because of the rigorousness of structure to function, such mutations usually lead to inactivity. They may be iso-proteins, but not iso-enzymes. But there are exceptions, two of which will be gotten into later.
              USEFUL VALUE. We would expect that within diploid organisms every enzyme would have its allele-mate. Hence, we would expect that any mutations of type "a", above would show up in electrophoresis even though the organism seems perfectly normal with regard to that particular function. All of Mendel's laws ought to be obeyed such as having the sum of individuals that are homozygous equal to those that are heterozygous. However, that is not the case in most instances, because within the population there are perhaps many different types of fully functional isozymes. Perhaps you have already studied the lac-operon. In the world's vast population of the mere bacterial species of E.coli, there are more than 15 electromorphs of &beta-galactosidase.

              Applying this to higher organisms, suppose you compared the electromorphs for an enzyme in earthworms taken from three locations: South Africa, Spain, and Hawaii. Suppose you found that the electrophoresis patterns between worms from South Africa and Spain seemed much more similar than to those in Hawaii. This would tell you that Hawaii's population had not had much genetic mixing with the world's other worms. Or suppose you did the same with white rhinos in various zoos and found that they were quite much more alike among themselves than they were with wild rhinos. That would tell you that zoo-bred rhinos were dangerously inbred, and that some new genes had to be brought in - more likely in the form of sperm taken from drugged wild rhinos.

              Let's look at a few more examples: compare and contrast the electromorphs of worms and cottonwood trees on both sides of a river that has flowed for at least the last 5,000 years. Or-, do the foxes in the western United States demonstrate much genetic mixing with those of the eastern United States? (A favorite haunt of foxes seems to be the median strips of Interstate Highways - culverts to live in and road kill and litter to forage.) Now do you see the power of this technique to the study of field biology, ecology, and endangered species?



              --------------------------------------------------------------------------------

              Doing Enzyme Electrophoresis

              One of the first things you need to do is make a starch gel slab. (See the associated page on materials and methods for doing this.)


              Once the slab is cooled and solidified, make a clean cut across the slab as shown here, and then pull the cut-off end away from the main portion a few inches.


              Cut small rectangles out of filter paper. Their widths should be about 1/4 inch (5 mm), and their lengths should be a little more than the depth of the starch gel you have made. On the bottom portion of the "wick" apply some of the mashed plant (or squash the plant right onto the wick. Do NOT allow it to dry (the enzymes will inactivate if dried.)


              Using a tweezers, lift the wick and stick it to the newly opened face of the starch gel. Make sure that the enzyme laden portion of the wick is stuck to the gel. Usually the first or last wick applied is one carrying the tracking dye - usually bromphenol blue (BPB or "buffalo black") BPB is a small negatively charged molecule that will move faster than any enzyme can move.


              After applying several wicks to the end of the gel - evenly spaced of course, and no closer to the edges than about 3/4 inch (15 mm), push the small piece of gel up against the wicks. In the diagram you can see one wick's tab protruding above the surface of the gel. (It used to be in the "old days" that scientists would cut little slits into the gel and then insert the wicks. The new way is much easier and less messy.


              Insert the gel into the electrophoresis chamber with the orientation shown. (Black is negative and red is positive when you are connecting the wire leads.) Run enzymes slowly so as to minimize heating, which denatures enzymes. continue running until the tracking dye is about 1 cm from the end of the gel near the positive pole. Then disconnect the power.


              Unseen in the gel are hundreds of different enzymes, To visualize the placement of specific enzymes, a number of tricks must be employed. In the case exhibited here, the desired enzyme is LDH (lactic dehydrogenase), which requires a coenzyme NAD+/NADH. The idea is that if the gel is soaked in a buffer containing lactate and NAD+, any place that LDH exists will see the oxidation of lactate → pyruvate with the reduction of NAD+ → NADH. Here comes the trick: the resulting NADH is used, in turn, to reduce a colorless compound that becomes colored (colorless → colored). Reiterating: as lactate goes to pyruvate, NADH is formed, which then converts the dye to a colored form. Here those reactions are shown figuratively:



              So we carefully slide the gel out of its tray into a container containing buffer with lactate and NAD+, let it gently slosh back and forth for awhile. Voila! Slowly color begins to develop in little bands wherever LDH had moved out of the plant juices. In this diagram, each of the three samples gave rise to two LDH bands, as might be expected of a diploid organism with two different alleles (heterozygous).


              Now here comes a really neat thing about working with enzymes that require the coenzyme NAD+: you can use the same buffer system as with LDH, but you just add a different substrate.



              But to do this you might think you need to make more gels, and thus need more equipment, BUT you don't! Just imagine that you were able to take an electrophoresed gel prior to "staining", and shave off thin sheets from the slab. Then treat each sheet with its own buffer. This can be done because starch gels are rather soft and "cuttable".



              Using the above device,

              First slip your gel into a tray that has lips of three sides. The one on the short side is called the stop, and the ones on the long sides are the cutting guides for the proper depth (it is very important that the top edges of this guide are smooth with no snags).
              Once the gel is in place, it is good to lay a flat piece of plexiglas or glass on top of the gel to hold it down and prevent it from buckling during the cutting process.
              Take a piece of smooth fishing line in both hands and, holding it very taut, draw it through the gel using the upper edges of the cutting guides to keep the depth of the cut constant.
              Once the cut has been made, remove the "hold-down plate" and gently slide the upper lay of gel onto that plate (it is good to have the plate wet as it makes sliding easier).
              Slide the bottom piece of gel out the end of the bottom frame into one of the staining containers containing buffer.
              Lay the top gel back into the bottom frame and repeat the cutting procedure.
              Do this until there is nothing left to cut.
              You now have several staining containers with gels floating around in them.
              Add the appropriate substrates and coenzymes to the various containers and start them gently oscillating to help mix the ingredients and expose all surfaces of the gel to the solution for even penetration of the chemicals into the gels.


              --------------------------------------------------------------------------------


              DISCUSSION

              There are two classic examples of proteins that are both iso-proteins and electromorphs. In the one case we have isozymes of LDH, which might be a surprise, but it became a classic because it was used years ago as a test for whether or not a person was having a heart attack. The other case is also connected with disease and deals with the iso-proteins of hemoglobin, and the disease is sickle-cell anemia - or is it a not a disease but rather a selective advantage in fighting another disease, malaria? 'Tis a strange world out there! Let us take a closer look.

              LACTIC DEHYROGENASE. In each of our bodies, we have genes for making two types of LDH, but those genes are not really alleles because alleles should theoretically come under the same controls that would turn them on and off. Also these two enzymes do something that any self-respecting catalyst should not do - alter the equilibrium. In striated muscles, one type is made, and, believe it or not, it is called "muscle-LDH"; and in the heart muscle there is a form called - you guessed it! - "heart-LDH". Their equilibria differ because their jobs are different. Under anaerobic exercise, muscles cannot shunt highly toxic pyruvate to the overloaded and aerobic Kreb's Cycle, so LDHM (muscle-LDH), which is a very fast enzyme, converts the pyruvate anaerobically to much less toxic lactate. The equilibrium is strongly in favor of lactate.

              On the otherhand, the highly aerobic heart muscle MUST not let moderately toxic lactate build up. So what it does is use LDHH (heart-LDH) to take any lactate that happens to be around, converts it to pyruvate and then evicts the pyruvate into the blood stream in hopes that somewhere else in the body can take care of it. The equilibrium is strongly towards pyruvate.

              Note that these are electromorphs as they move to different places in a gel. Normally your blood contains a fair amount of LDHM because our movements squish a few muscle cells here and there releasing this enzyme into the blood stream. But a healthy heart keeps all its LDHH to itself - UNLESS a heart attack damages the tissue and releases that enzyme into the blood stream. So, in the old days, if your blood gave rise to two bands of LDH activity, it meant that you had had a heart attack. (Other tricks were also used to amplify the effect making use of the fact that both LDH's are really tetramers in the quaternary structures. Make your teacher show you how if the heart attack victim's two types of LDH's were dissociated into monomers and then recombined, the electrophoresis would result in FIVE bands, which leap out at the technician to say: "That person needs help quickly!" (The test was too slow - 20 minutes - and a faster test of a coronary infarction had to be found to save even more lives.)

              HEMOGLOBIN. Most of use carry two identical alleles for "normal" hemoglobin. Were you to electrophorese your blood, you would get only one red band in the gel. (Note that staining is not needed.) However a few percentage of the black population would show up with two red bands. If you took their blood and deoxygenated it, you would see a large proportion of their erythrocytes collapse and form crescents like the farm tool called a sickle. And a very few of that population would have a single red band that was different from the "normal" band found in most of the human population. In other words, most people are homozygous "normal", a few are heterozygous normal/sickle, and a very few are homozygous sickle. The last often don't live very long. What is more is that there are four electromorphs of "sickle" found in the world. There is a major type widely spread in its African origin - and that indicates it came into being thousands of years ago, and because it persists it must have conferred upon its carriers some sort of advantage. Indeed, those people are not affected severely by malaria that is rampant in the tropics. The other three forms have arise more recently in West Africa, and form small pockets on the map. Those people also are resistant to malaria.

              Thus comes the question: is sickle cell anemia a positive or a negative? Were it not for modern medicine, it would be a positive as it helps people to survive a malaria infected world. But with modern medicine's combatting malaria it is a negative, and is one of those diseases caused by advancing culture. Want three more "cultural advancement" diseases? Polio, menopause, and cancer. In the "olden days" most people didn't live long enough to encounter the last two, and the first one everyone got as a newborn entering a squalid environment. At that age the paralytic attribute cannot manifest itself, and the disease is merely a runny nose for a few days. (The major motor nerves must be myelonated to have polio myelitis, and pre-toddlers don't have myelonated nerves yet - that's why they are so uncoordinated.)

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              • #8
                http://www.vivo.colostate.edu/hbooks...rinciples.html


                http://www.dnalc.org/ddnalc/resources/animations.html
                http://www.sumanasinc.com/webcontent/animation.html

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                • #9
                  شكرا
                  لك
                  طولتها وعرضتها
                  الموضوع أبسط من كذا بكثير

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                  • #10
                    فيه مواضيع بالمنتدى عن Electrophorsis

                    http://www.arabslab.com/vb/showthread.php?t=639

                    http://www.arabslab.com/vb/showthread.php?t=2933

                    http://www.arabslab.com/vb/showthread.php?t=657

                    http://www.arabslab.com/vb/showthread.php?t=754

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                    • #11
                      السلام عليكم ورحمة الله وبركاته
                      أعذروني انا دارس في سوريا باللغة العربية وسوف أشرح باللغة العربية واعذورني من نسيان أسماء بعض المواد فأنني لم أجري التحليل منذ اكثر من أربع سنوات مع انني لي باع طويل فيه وخبرة مميزة والحمد لله مع العلم انه تحليل في غاية الدقة والحساسية .
                      هناك أكثر من نوع للرحلان فهناك رحلان بروتين - رحلان خضاب - رحلان شحوم - رحلان سائل دماغي شوكي - رحلان بول - رحلان كولسترول - رحلا LDH
                      والأكثر شيوعاً هي رحلان الخضاب ورحلان البروتين
                      رحلان الخضاب يفيد في تشخيص حلات فقر الدم الأساسي فهو يحدد نوع الخضاب إن كان A1 - F -S -A2 كما وهناك بعض الأنواع النادرة جدا
                      ووجود النوع ِA1 بنسبة 95 إلى 100 % هو الحالة الطبيعية للأنسان السليم وذلك بعد العام الأول من عمر الانسان ( حيث وجود الخضاب F فبل عمر السنة يعنبر طبيعي طبعا حسب النسبة فالخضاب F هو المسمى الخضاب والوليدي في يكون في الأيام الاول ربما ألى 98 % ويبدأ في التناقص مع تقدم العمر حتى يصل إلى 0 % ووجود الخضاب F بعد سن العام الاول يدل على أن المريض مصاب بالثلاسيميا ( بوجد اكثر نوع من الثلاسيما صغرى وكبرى ومتوسطة يطول شرحها ربما في وقت آخر ) ووجود الخضاب s يدل على الاصابة بالتمنجل
                      ويجر عادة تحليل رحلان الخضاب عادة على دم مأخوذ على ال EDTA
                      حيث تقوم أولا بعملية حل للدم في مادة مخصصة
                      ومن ثم تطبيقها على على شرائح سللوزية مخصصة ( بعد نقع هذه الشرائح في مادة خاصة بها ) ومن تم وضع االشريحة على جهاز الترحيل /ن القطب الموجب إلى السالب حيث يأخذ كل خضاب مكانه المناسب بعد حوالي 20 دقيقة تختلف بحسب شدة الفولتاج وذلك وفقاً لثقلة النوعي وبعد ذالك نلون الشريحة بملون مناسب لها ونقوم بتثبيته وإظهاره حتى تظهر الشريحة بلون شفاف نقوم بتنشيف الشريحة ثم القرآة على جهاز الرحلان علما أنك بإستطاعتك التشخيص من خلال الشريحة فوراً ولكن من الواجد القرآة على الجهاز حتى تستطيع ارسال التحليل بالشكل المناسب وللمعوم فكل خضاب يأخذ مكانه المناسب ولا يتعداه
                      وسوف اكمل شرح رحلان البروتين ورحلان الشحوم في وقت سابق ولكني أريد رأيكم هل الشرح كافٍ ام ماذا وهل هناك استفسارت أخرى علما ان التحليل شرحه بشكل عملي سهل جداص ولكن سامحوني طريقة شرحي بسيطة وارتجالية وليست محضرة من قبل

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                      • #12
                        يعطيك العافيه ابو البراء شرح بسيط ومفهوم..
                        sigpic

                        قال صلى الله عليه وسلم:
                        (ثلاث كفارات وثلاث درجات وثلاث منجيات وثلاث مهلكات فأما الكفارات فإسباغ الوضوء في السبرات وانتظار الصلوات بعد الصلوات ونقل الأقدام إلى الجماعات وأما الدرجات فإطعام الطعام وإفشاء السلام والصلاة بالليل والناس نيام وأما المنجيات فالعدل في الغضب والرضا والقصد في الفقر والغنى وخشية الله في السر والعلانية وأما المهلكات فشح مطاع وهوى متبع وإعجاب المرء بنفسه)

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                        • #13
                          السلام عليكم الموضوع بسيط وشيق جدا باختصار مثلا فى شغل
                          extraction of dna
                          دا بيكون خطوه اساسيه هو الelectrophoresis

                          . the stander method used to seprate identify purify dna fragment through agrose gels the location of dna with in gel determined directly bands of dna in gel stained with intercalting dye
                          لو حد محتاج التفاصيل وخطوات العمل ممكن انزلها فى الموقع
                          وربنا يوفق الجميع:sm199:

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                          • #14
                            السلام عليكم
                            انا عندي موقع ممتاز ولكن باللغه الالمانيه
                            الذي لاعرف الالماني ممكن انه يشاهد الصور
                            لانه ممكن بالصور بفهم الطالب حيث يحتوي الموقع على صور تفيد كيفيه العمل
                            http://www.med4you.at/laborbefunde/t...htm#Einleitung
                            ان شاء الله بالتوفيق للجميع

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                            • #15
                              electrophorsis : هي طريقة تستعمل لفصل الاجزاءالبروتينة المكونة للبروتين الكلي في مصل الدم
                              sepearation of protein fractions composing total protein in serum


                              وتستند الفكرة الاساسية في عملية الفصل على اعتبار البروتينات جزيئات تحمل شحنة كهربائية سالبة اذا ما وضعت في محلول قاعدي

                              وعلى هذا الاساس فإن الاجزاء البروتينية على اختلاف مكوناتها وانواعها تتصرف كايونات سالبة اذا ما مرر تيار كهربائي فيها ولذلك فهي تنجذب نحو القطب الموجب ANODE وان سرعتها الحركية تعتمد على كثافة الشحنة الكهربائية على سطح الجزيئة البروتينية بصورة رئيسية وعلى عوامل اخرى مثل شدة التيار الكهربائي المار والتركيز الهيدروجيني للمحلول الدارئ اضافة الى الوزن الجزيئي للجزء البروتيني المنفصل والوسط الداعم supporting medium المستخدم في عملية الفصل .... هذه فكرة اساسية عن الموضوع
                              [ THEORY WITHOUT PRACTICE IS EMPTY ...PRACTICE WITHOUT THEORY IS BLIND[/ ]

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