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Analytical Techniques Good Laboratory Practice 08.April 2013

International Max Planck Research School for Global Biogeochemical Cycles. Analytical Techniques Good Laboratory Practice 08.April 2013. Ines Hilke Iris Kuhlmann. content Introduction Please introduce yourself and briefly your project

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Analytical Techniques Good Laboratory Practice 08.April 2013

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  1. International Max Planck Research School for • Global Biogeochemical Cycles Analytical TechniquesGood Laboratory Practice08.April 2013 • Ines Hilke • Iris Kuhlmann

  2. content • Introduction • Please introduce yourself and briefly your project • What do you expect from this today’s training course and give one sentence or a few words – • what does GLP “Good Laboratory Practice” represent for you • The aim of our day • Good Laboratory Practice – what's that and the need? • Quality management systems • Troubles course (make four groups) • Good Laboratory Practice • Qualification, Validation, System Suitability Test • Documentation • Weighing and pipetting is one of the most frequent jobs done in the lab • How much is a Gram • Analytical balance and electromagnetic force compensation • Metric systems and traceability • How to install the balance and using an Analytical Balance • Calibration and calibration certificate • Uncertainty of your balance • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  3. International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 ‘Chapter’ - type here

  4. Management Requirements and Technical Requirements. Management requirements are primarily related to the operation and effectiveness of the quality management system within the laboratory. Technical requirements includes factors which determines the correctness and reliability of the tests and calibrations performed in laboratory. Laboratories use ISO/IEC 17025 to implement a quality system aimed at improving their ability to consistently produce valid results.[2] It is also the basis for accreditation from an accreditation body. Since the standard is about competence, accreditation is simply formal recognition of a demonstration of that competence. A prerequisite for a laboratory to become accredited is to have a documented quality management system. The usual contents of the quality manual follow the outline of the ISO/IEC 17025 standard. WIKI • International Max Planck Research School for • Global Biogeochemical Cycles Event name, date

  5. The aim of the day • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  6. The aim of the day Today we will have look at some tools that could help you: • avoid being trapped in this cog wheel, • secure constant high quality of your work today and in future, • provide excellent results in time, • be aware of errors • and make sure you are able to • find them easily. • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  7. The aim of the day GLP Good Laboratory Practice – what's that and why the need? • In the beginning GLP intended for non-clinical studies conducted for the assessment of the safety or efficacy of environmental dangerous chemicals and pharmaceuticals to man, animals and the environment. AND it was designed • To improve Lab standards and to ensure uniformity, consistency, reliability, reproducibility, quality, and integrity of results and to ensure that anyone can fully reconstruct the results. • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  8. The aim of the day TODAY GLP refersto a quality system of management controls for research laboratories (non-clinical) and organisations An internationally recognized definition of GLP can be found on the website for the Medicines and Healthcare products Regulatory Agency-UK which defines GLP as: • Good Laboratory Practice (GLP) embodies a set of principles that provides a framework within which laboratory studies are planned, performed, monitored, recorded, reported and archived. These studies are undertaken to generate data by which the hazards and risks to users, consumers and third parties, including the environment, can be assessed for pharmaceuticals (only preclinical studies), agrochemicals, cosmetics, food additives, feed additives and contaminants, novel foods, biocides, detergents etc.... GLP helps assure regulatory authorities that the data submitted are a true reflection of the results obtained during the study and can therefore be relied upon when making risk/safety assessments. • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  9. The aim of the day QM - Quality management systemshave more or less similar requirements Conclude: • with QM protocols you achieve that procedures are transparent and traceable • errors can be easily found • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  10. Troubles course Troubles course Topic A (2 groups) How would you proof the reliability of an laboratory, e.g. Environmental Analysis, as an inspector of the Ministry for Social Affairs? • identify the hot spots in the lab and • discuss the recognizable issues • take some notes to be able to discuss the measures with the head of the laboratory • position your questions cards at the hot spots • do you have any further suggestions? Topic B (2 groups) The laboratory of Environmental Analysis is blamed for “false alarm” an your are the head and you are facing a lawsuit and a high fine. Please make a quality control check and inform yourself about GLP to be strengthened against allegations. • are there any critical issues in the lab? Can you identify the hot spots? • how would you check the traceability of the obtained results? • please take some notes to be able to discuss the measures with your staff • position your cards at the hot spots • do you have any further suggestions? • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  11. Break • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  12. GLP Good Laboratory Practice GLP Good Laboratory Practice Project or units of analysis Result and information conclusion Field trip and sampling Sample preparation measurement evaluation Basic principle Analytical method Analytical procedure • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  13. GLP Good Laboratory Practice GLP Good Laboratory Practice Example: You would like to buy a new balance or measurement device, or you would like to change a method procedure. 3 Steps to go… • you have to check whether the machine is suitable for the purpose you would like to buy it for. Proof of suitability or Qualification of the device • check the measurement method – Validation • test if method and machine achieve the expected attainment - System Suitability Test • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  14. GLP Good Laboratory Practice • Step 1. Proof of suitability or Qualification of the device Project or units of analysis Result and information conclusion Field trip andsampling Sample preparation measurement evaluation Basic principle Analytical method Analytical procedure • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  15. GLP Good Laboratory Practice • Step 1. Proof of suitability or Qualification of the device • Check if ... • equipment installed under proper conditions, operates properly, • accuracy & precision e.g. with a service contract, calibration service consumer’s job - “product requirement document” vendor’s job - “functional specification” • Prove that the equipment consistently maintains a certain level of performance when is used for the measurement of samples. • Assure that the equipment consistently provides reliable and accurate measurements in daily analyses • Check if... • delivered equipment is in accordance with the requested design and specifications, • equipment has been installed under proper conditions, • environment is adequate for the equipment. • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  16. GLP Good Laboratory Practice Accuracy & precision • Step 1. Proof of suitability or Qualification of the device The accuracy of a measurement system is the degree of correctness of measurements of a quantity to that quantity’s actual (true) value accuracy Systematic errors excellent bad accurate & precise NOT accurate precise but precision Random errors Neither accurate or precise NOT precise but accurate bad The precision of the measurement system, also called reproducibility or repeatability, is the degree to which repeated measurements under unchanged conditions show the same results • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  17. GLP Good Laboratory Practice GLP Good Laboratory Practice Example: You would like to buy a new balance or measurement device, or you would like to change a method procedure. 3 Steps to go • you have to check whether the machine is suitable for the purpose you would like to buy it for. Proof of suitability or Qualification of the device • check the measurement method – Validation • test if method and machine achieve the expected attainment - System Suitability Test • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  18. GLP Good Laboratory Practice • Step 2. Validation Project or units of analysis Result and information conclusion Field trip and sampling Sample preparation measurement evaluation Basic principle Analytical method Analytical procedure • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  19. GLP Good Laboratory Practice • Step 2. Validation validation is the documented act of demonstrating that a procedure, process, and activity will consistently lead to the expected results. The validation of the analytical method follows the same principles like the qualification. Now the principles are related to the performance of the analytical method • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  20. GLP Good Laboratory Practice • Step 2. Validation validation is the documented act of demonstrating that a procedure, process, and activity will consistently lead to the expected results. The validation of the analytical method follows the same principles like the qualification. Now the principles are related to the performance of the analytical method • Besides • accuracy & precision • Additional • systematic deviations • limit of quantification/determination “LOQ” • linearity • proof of the following parameters is recommended • selectivity & specificity • limit of detection “LOD” • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  21. GLP Good Laboratory Practice GLP Good Laboratory Practice Example: You would like to buy a new balance or measurement device, or you would like to change a method procedure. 3 Steps to go • you have to check whether the machine is suitable for the purpose you would like to buy it for. Proof of suitability or Qualification of the device • check the measurement method – Validation • test if method and machine achieve the expected attainment – System Suitability Test • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  22. GLP Good Laboratory Practice GLP Good Laboratory Practice Project or units of analysis Result and information conclusion Field trip and sampling Sample preparation measurement evaluation Basic principle Analytical method Analytical procedure • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  23. GLP Good Laboratory Practice • Step 3. SST System Suitability Test The SST could be part of the method validation and it checks if method and machine achieve the expected attainment. 2 steps to go.. 1. functioning of the measurement device, 2. check the measurement with repeated tests during the routine. • Laboratory Information Management System (LIMS) • A feature, either ready to buy as soft ware package or self-made. The design is variable, depending on your needs. • For example it could support: • data tracking of all measurement results, • data exchange interfaces, • helps with data evaluation • includes a Quality Control Chart • e.g. Minerva/balance – self-made • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  24. GLP Good Laboratory Practice Documentation… One more Step to go … Documentation of all …your actions • SOP (Standard Operation Procedure) • Quality Control Charts for each method • “log book” of each device, documentation of the daily performance, via paper or electronically • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  25. GLP Good Laboratory Practice Documentation… One more Step to go … Documentation of all …your actions Documentation has to be transparent, reproducible and traceable Rules for documentation: • something that isn't documented, was not done - official • “W-rule” Who has done What, When, With What equipment and Why. • Compulsory archiving – 15 years. MPI Rules of “Good Scientific Practice” “….reliable securing and storage of primary data for 10 years clear and comprehensible documentation of methods (e.g. lab book) and all important results…” • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  26. GLP Good Laboratory Practice Documentation… Documentation has to be transparent, reproducible and traceable NO ... 6 7 3 overstrike of first number – first number is not visible anymore 6 7 Tipp ex – fist number is not visible anymore 6 7 3 5 first number not visible anymore 6 7 3 5 No reason for this correction mentioned • Correct! • 6 7 3 5 5 01/09/2012 IK Reading error • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  27. GLP Good Laboratory Practice Documentation … Documentation has to be transparent, reproducible and traceable • NO ... • Correct! • You sample bags should be labelled with the ? F 13 F 15 F 14 F 13 F 16 Sample date Plot name or number Contributor or project name • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  28. Break • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  29. Balance Weighing and pipetting is one of the most frequent jobs done in the lab The balance (also balance scale, beam balance and laboratory balance) was the first mass measuring instrument invented.[11] Sanders, L. A Short History of Weighing. Birmingham, England: W. & T. Avery, Ltd. (1947, revised 1960) Wikipedia A balance compares weights, not masses, the weight of an object is proportional to its mass, and the standard weights used with balances are usually labeled in mass units. (WIKI) How much is a GRAM 1 t = 1000 kg 1 kg = 1000 g 1g = 1000 mg 1 mg = 1000 µg • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  30. Balance How much is a GRAM Digit reading precision Type of balance 0 1 2 3 4 5 6 7 1 g 0,1 g 0,01 g 0,001 g 0,0001g 0,00001 g 0,000001 g 0,0000001 g 1/10 g 1/100 g 1 mg 1/10 mg 1/100 mg 1 µg 1/10 µg Precision balance Precision balance Precision balance Analytical balance Semi-micro balance Micro balance • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  31. Balance TODAY Electronic analytical “balance” Electronic analytical scale measures the force needed to counter the mass . • They use an electromagnet to generate a force to counter the sample being measured and outputs the result by measuring the force needed to achieve balance. Such measurement device is called electromagnetic force compensation. • This makes calling it an "analytical balance“ an unsuitable name, because it should actually be called an "analytical scale", due to it measuring force, rather than gravitational mass. • As such they must have calibration adjustments made to compensate gravitational differences. • The electromagnetic force compensation is a excellent measurement principle for electronic scales in the highest accuracy class. • It is designed to measure small mass in the sub-milligram range. • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  32. Balance Electromagneticforcecompensation Operation principle Withthis principle, the loads weight is compensated by an electro magnetically generated force. A compensation coil through which a permanent current flows is inserted in a permanent magnetic field. In the unloaded condition, current regulation ensures that the system is in the zero position. With the aid of an opt electronic position sensor, the coil position is controlled to an accuracy better than a thousandth of a millimeter. It records vertical positional changes when the scale is loaded. This information from the controller is used to generate a compensation current in the coil that returns the weight system to zero. The current is directly proportional to the loaded weight. Its value is digitalized and sent to the display. Coil Force Picture: Mettler Toledo Picture: Mettler Toledo • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  33. Balance Metric systems and traceability • ……. The first practical realization of the metric system came during the French Revolution, when the existing system of measure which had fallen into disrepute was replaced by a decimal system based on the kilogram and the metre. • Until 1875, the French government owned the prototype metre and kilogram • 1875 the CONVENTION OF THE METRE was signed, and the control of the standards relating to mass and length passed to a trio of inter-governmental organisations. • The International Bureau of Weights and Measures (Bureau international des poids et mesures or BIPM) was born  – an organisation based at Sèvres, France that supervises the International Prototype of the KILOGRAM, and provides metrology services for the GCPM (Conférence générale des poids et mesures)and CIPM (Comité international des poids et mesures) • In 1960, the CGPM launched the International System of Units (in French the Système international d'unités or SI) which had six base units, the metre, kilogram, second, ampere, degree, kelvin and 22 derived units. • The metre was retained as the international standard until 1960 when the metre was redefined in terms of the wavelength of the orange-red line of kryptonkryp-86. • As of 2012 the prototype kilogram was still in use, though it is expected to be replaced by a new definition within the next few years. • Eventually every kilogram is must be traceable back to the national and/or international standard kilogram • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  34. Balance Things you should know before using a balance • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  35. Balance How to install the balance • stable table, suitable for an analytical balance • stable and consistent laboratory conditions • install the balance where the influence of the air current is insignificant / low • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  36. Balance How to install the balance • stable table, suitable for an analytical balance to avoid vibration • install the balance on an stable anti-vibration table made out of stone • the table has to be non magnetic – no steel plate • the table has to be protected against electrostatic charging (NO plastic or glass) • make sure the anti-vibration table is only fixed once – either on the floor or at the wall • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  37. Balance How to install the balance • Stable and consistent laboratory conditions • make sure the anti-vibration table is put up in an suitable laboratory • without vibration – in the corner of room is the most stabilized place within the building • prefer consistent laboratory conditions. Do not place the balance where there could be rapid temperature change ( e.g. near a window with direct sunlight, heater or lamp) • air currents - avoid an location near air conditioner, near a doorway, passageway, wherever there is a temperature change • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  38. Balance • BAD or NO reproducibility of results • NO stability • TO slow • unclean and grubby balance • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  39. Balance Using an Analytical Balance • Air currents (Wind) and unsteady measurements Air current influence are observable: • weighing becomes unstable • Measures to counter • The measuring pan of an analytical balance (> 0.1 mg ) is inside a transparent enclosure/ draft shield with doors so that air currents do not affect the balance's operation. • close the enclosure during weighing operation • open the enclosure just as far as necessary. Picture: Mettler Toledo • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013

  40. Balance Using an Analytical Balance • Static electricity and unsteady measurements • Static electricity influence is observable: • slow drift of the weighing result in one direction • poor reproducibility in repeated weighing procedures Picture: Mettler Toledo • Causes of Static Electricity • Frictional electrification occurs when objects are rubbed together. • The electrostatically charged samples and the objects surrounding the sample attract each other. • e.g. through: • pouring liquid or powder from one container to another • toughing glass with gloves - nonconductive containers (glass or plastic) are electrically charged. Then electrostatically charged flasks and the objects surrounding the flask/sample attract each other • wearing synthetic underwear • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  41. Balance Using an Analytical Balance • Static electricity and unsteady measurements • Measures to counter: • maintaining sufficient air humanity > 45 - 60 % • ionize the air in and around the balance with an ionizing blower • don’t use plastic containers • shield the sample with conductive enclosure e.g. “ErgoClips” Picture: Mettler Toledo • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  42. Balance Using an Analytical Balance Magnetic force und unsteady measurements • Magnetic force is observable: • poor reproducibility in repeated weighing procedures and • depending of the flask position on the weighing pan • result stays stabile Causes of magnetic force When measuring a magnetic material it may attract and be attracted to the surrounding components of the balance, or be influenced by the magnetic force of the sensor coil. This will lead to inaccurate weighing results. • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  43. Balance Using an Analytical Balance Magnetic force und unsteady measurements • Measures to counter magnetic force: • take the magnetic stirrer out of the liquid • increase a distance between the sample and the balance and use the underhook • shield the sample and use an mu-metall or permalloy foil or • take other weighing container or beaker glass • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  44. Balance Using an Analytical Balance Convection flow and unsteady measurements • Convection flow is observable: • drift of the weighing result in one direction • sample that are hotter appear lighter and those that are colder, heavier • Causes of convection flow: • Convectionof air from cold items - balance is not acclimated • or hot items – sample container is to hot • Condensation of atmospheric water on cold items/sample - sample gain weight • Evaporationof water from wet items/sample - sample loose weight Picture: Mettler Toledo • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  45. Balance Using an Analytical Balance • Convection flow and unsteady measurements • Measures to counter convection flow: • acclimation of the balance - leave the balance always in the stand by mode • the sample and the balance must be at room or the same temperature to prevent natural convection • refrain from directly touching the sample or the tare – use tweezers instead • keep you sample flask closed or use narrow neck flasks, • use dry weighing container • mark your flask before hand in time – solvents evaporating, • use “Fact” - Fully Automatic Calibration Technology • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  46. Balance Using an Analytical Balance Gravity and unsteady measurements • Gravity is observable: • reproducibility in repeated weighing procedures are poor when moving the balance to a different height or over a large distance • Gravitational force • the electronic balance measures the gravitational force on the sample. Therefore, if the acceleration of gravity varies, the display value will also vary. • objects on the earth are constantly subjected to the “universal gravity” which attracts them towards the centre of the earth, as well as the “centrifugal force” caused by the rotation of the earth • the acceleration of gravity is not the same everywhere on earth Picture: Mettler Toledo • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  47. Balance Using an Analytical Balance • Acceleration of gravity and unsteady measurements • Moving the analytical balance in height The measured value will change when the weighing instrument is moved to a different height. When the weighing instrument is placed one floor higher (4 m), a weight of 200 g will weigh 0.26 mg less. • Moving the analytical balance over a larger distance • The acceleration of gravity varies with latitude and altitude. • As a consequence of centrifugal forces caused by the earth’s rotation it is less at the equator than at the poles. • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  48. Using an Analytical Balance Effects of tilting and the acceleration of gravity and unsteady measurements When a balance is placed horizontally, it detects the gravitational force that is applied to the sample vertically. Gravitational force = Display value • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  49. Balance Using an Analytical Balance Effects of tilting and the acceleration of ravity and unsteady measurements Non- horizontal position of the weighing instrument If the balance is placed on a slope, the gravitational force applied to the Sample will be broken down to the longitudinal and lateral directions of the balance. As the balance detects only the longitudinal force, it will display a smaller value than the actual weight of the sample. If the balance is moved to a table that is not leveled, but is tilted 5 mm on a length of 1m deviates 0.2865° from the reference position. That means a weight of 200 g has an 2.5 mg lower readout. The zero setting also changes: the zero point is also a weight value, it is the weight of the empty weighing pan. The force that cannot be detected Display value Gravitational force • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

  50. Balance Using an Analytical Balance Effects of tilting and the acceleration of gravity and unsteady measurements • Measures to counter the effects of tilting and the acceleration of Gravity: • after each relocation you have to do a calibration and readjustment the balance • level the balance using its levelling feet • ff levelling is difficult, calibrate the balance in the current position • International Max Planck Research School for • Global Biogeochemical Cycles Good Laboratory Practice 08.April 2013 Event name, date

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