Deuterium pulse width calibration and decoupling - Revision history

Alex: /* 13C Anti-Phase Triplet - Indirect Calibration */

2010-01-05T23:04:27Z

13C Anti-Phase Triplet - Indirect Calibration

Alex: /* 13C In-Phase Triplet */

2010-01-05T22:59:48Z

13C In-Phase Triplet

← Older revision		Revision as of 22:59, 5 January 2010
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	At this step you will need the <tt>H2_pwxcal</tt> dataset. If you don't have this dataset, install it:		At this step you will need the <tt>H2_pwxcal</tt> dataset. If you don't have this dataset, install it:

	#Downloading [[~~NESG~~:~~%ATTACHURL%/H2 pwxcal.tar.gz\|~~H2_pwxcal.tar.gz]] into your VNMRJ user home directory		#Downloading [[File:H2_pwxcal.tar.gz]] into your VNMRJ user home directory
	#Ucompress and unpack it		#Ucompress and unpack it
	#Go to <tt>~/vnmrsys/psglib/</tt> and compile the pulse sequence with <tt>seqgen H2_pwxcal.c</tt>		#Go to <tt>~/vnmrsys/psglib/</tt> and compile the pulse sequence with <tt>seqgen H2_pwxcal.c</tt>
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	#Using the current dataset calibrate the 13C 90-degree pulse with the 360 method, along the lines described for 1H (1H cal).<br>Adjust <tt>tpwr</tt> so that <tt>pw</tt> is equal or greater then the typical <tt>pwC</tt> pulse.		#Using the current dataset calibrate the 13C 90-degree pulse with the 360 method, along the lines described for 1H (1H cal).<br>Adjust <tt>tpwr</tt> so that <tt>pw</tt> is equal or greater then the typical <tt>pwC</tt> pulse.

	Please note that 13C T1 relaxation is very slow, and its effect on the accuracy of 13C pulse width calibration is even stronger than for 1H.		Please note that 13C T1 relaxation is very slow, and its effect on the accuracy of 13C pulse width calibration is even stronger than for 1H.



	==== 13C Anti-Phase Triplet - Indirect Calibration ====		==== 13C Anti-Phase Triplet - Indirect Calibration ====

Alex: /* 2H 90° Pulse - Manual Calibration */

2010-01-05T22:46:35Z

2H 90° Pulse - Manual Calibration

← Older revision		Revision as of 22:46, 5 January 2010
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	#Tune the <sup>13</sup>C and <sup>2</sup>H channels as described on the [[Tuning_and_matching\|tuning page]].		#Tune the <sup>13</sup>C and <sup>2</sup>H channels as described on the [[Tuning_and_matching\|tuning page]].
	#Lock the sample on the benzene-d6 signal; <tt>z0</tt> will be different from that when locking on D<sub>2</sub>O!		#Lock the sample on the benzene-d6 signal; <tt>z0</tt> will be different from that when locking on D<sub>2</sub>O!
	#Shim the sample as described on the [[~~SampleShimming~~\|shimming page]]. You can use <sup>1</sup>H gradient shimming on the dioxane resonance.		#Shim the sample as described on the [[Shimming\|shimming page]]. You can use <sup>1</sup>H gradient shimming on the dioxane resonance.

	=== 1D Deuterium Spectrum ===		=== 1D Deuterium Spectrum ===
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	Also see the instructions given in <tt>man('BioPacklist')</tt>.		Also see the instructions given in <tt>man('BioPacklist')</tt>.

	For details on probefile update see [[NESG:BioPackProbefile#Manual_Probefile_Update\|Manual Probefile Update]] page.		For details on probefile update see [[NESG:BioPackProbefile#Manual_Probefile_Update\|Manual Probefile Update]] page.

	== Using 2H Decoupling With Deuterated Protein Samples ==		== Using 2H Decoupling With Deuterated Protein Samples ==

Alex at 22:45, 5 January 2010

2010-01-05T22:45:56Z

← Older revision		Revision as of 22:45, 5 January 2010
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	Pulsing on deuterium is most commonly used as deuterium decoupling in triple-resonance experiments with deuterated samples. Whenever aliphatic <sup>13</sup>C magnetization is transverse, it must be decoupled from the attached deuterons. Omitted or improperly calibrated 2H decoupling would cause a sharp sensitivity drop due to scalar relaxation of the second kind (~~insert reference here~~).		Pulsing on deuterium is most commonly used as deuterium decoupling in triple-resonance experiments with deuterated samples. Whenever aliphatic <sup>13</sup>C magnetization is transverse, it must be decoupled from the attached deuterons. Omitted or improperly calibrated <sup>2</sup>H decoupling would cause a sharp sensitivity drop due to scalar relaxation of the second kind <ref>Murali, N., Rao, B.D.N. (1996) Lineshape variations of a spin-1/2 nucleus coupled to a quadrupolar spin subjected to RF irradiation. ''JMR, Series A'', '''202'''(3), 215-221</ref>. Thus, proper application of <sup>2</sup>H decoupling requires tuning the 2H channel and calibrating <sup>2</sup>H 90° pulse.

	~~Proper setup requires tuning the 2H channel and calibrating 2H 90-degree pulse.~~		== 2H 90° Pulse - Automatic Calibration ==

	== 2H 90° Automatic Calibration ==

	See 'Chapter 9.0: '''2H AutoCalibration'''' in the BioPack manual.		See 'Chapter 9.0: '''2H AutoCalibration'''' in the BioPack manual.
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	The automatic calibration method of BioPack uses the same protein sample and calibrates the pulse directly on the HDO signal. It requires a K5022 relay to be present, which allows pulsing deuterium on the 4th channel, while detecting it. At SUNY Buffalo 600 MHz and 750 MHz instruments don't have this relay, therefore requiring manual calibration.		The automatic calibration method of BioPack uses the same protein sample and calibrates the pulse directly on the HDO signal. It requires a K5022 relay to be present, which allows pulsing deuterium on the 4th channel, while detecting it. At SUNY Buffalo 600 MHz and 750 MHz instruments don't have this relay, therefore requiring manual calibration.

	== 2H 90° Manual Calibration ==		== 2H 90° Pulse - Manual Calibration ==

	The current protocol is similar to the '''Calibration via Indirect 2H Detection using 13C Observe''' section of Chapter 9.0 '''2H AutoCalibration''' in theBioPack manual. Though you can still use the <tt>ddec_pwxcal</tt> dataset as recommended in the BioPack manual, the method presented here is more accurate and easier to use.		The current protocol is similar to the '''Calibration via Indirect 2H Detection using 13C Observe''' section of Chapter 9.0 '''2H AutoCalibration''' in theBioPack manual. Though you can still use the <tt>ddec_pwxcal</tt> dataset as recommended in the BioPack manual, the method presented here is more accurate and easier to use.

	Manual calibration of deuterium 90° pulse requires an ASTM standard test sample, which contains 60% benzene-d6 (~~C6D6~~) and 40% dioxane. The chemical shifts of benzene are 7.15 ppm ~~for 2H~~ and 128.0 ppm ~~for 13C~~ (http://www.sdsnmr.com/cs_table.html)~~; 1J_CD =~~ 24 Hz. Note that 2H and 1H shifts are essentially the same and that benzene-d6 contains ~~13C~~ at natural abundance level.		Manual calibration of deuterium 90° pulse requires an ASTM standard test sample, which contains 60% benzene-d6 (C<sub>6</sub>D<sub>6</sub>) and 40% dioxane. The chemical shifts of benzene are 7.15 ppm (<sup>2</sup>H) and 128.0 ppm <sup>13</sup>C (see http://www.sdsnmr.com/cs_table.html) and the scalar coupling <sup>1</sup>J<sub>CD</sub> is 24 Hz. Note that <sup>2</sup>H and <sup>1</sup>H shifts are essentially the same and that benzene-d6 contains <sup>13</sup>C at natural abundance level.

	#Insert the ASTM sample		#Insert the ASTM sample
	#Tune the ~~13C~~ and 2H channels as described on the [[~~NESG:RFCoilTuningAndMatching#13C_15N_and_2H_Channels_All_Prob~~\|tuning page]].		#Tune the <sup>13</sup>C and <sup>2</sup>H channels as described on the [[Tuning_and_matching\|tuning page]].
	#Lock the sample on the benzene-d6 signal; ~~'''~~z0~~'''~~ will be different from that when locking on ~~D2O~~!		#Lock the sample on the benzene-d6 signal; <tt>z0</tt> will be different from that when locking on D<sub>2</sub>O!
	#Shim the sample as described on the [[~~NESG:~~SampleShimming\|shimming page]]. You can use 1H gradient shimming on the dioxane resonance~~; set <tt>d1=10</tt> for best results~~.		#Shim the sample as described on the [[SampleShimming\|shimming page]]. You can use <sup>1</sup>H gradient shimming on the dioxane resonance.

	=== 1D Deuterium Spectrum ===		=== 1D Deuterium Spectrum ===

	Typical ~~2H 90-degree~~ pulse width for decoupling is ~~~280~~ us. With a pulse that long, it is important to know the exact 2H offset of benzene-d6 to ensure ~~the~~ on-resonance condition. You can use a 1D deuterium spectrum of the test sample to find the resonance. This experiment will also test if you can pulse and detect deuterium on the first channel.		Typical <sup>2</sup>H 90° pulse width for decoupling is in the order of 200-300 us. With a pulse that long, it is important to know the exact <sup>2</sup>H offset of benzene-d6 to ensure on-resonance condition during calibration. You can use a 1D deuterium spectrum of the test sample to find the resonance. This experiment will also test if you can pulse and detect deuterium on the first channel.

	The current protocol is based on the "Directly Observing Deuterium" in the [http://www.varianinc.com/image/vimage/docs/products/nmr/apps/pubs/manuals/0199914100a.pdf Deuterium Decoupling Channel Installation] manual with small modifications. To record a 1D deuterium spectrum do the following:		The current protocol is based on the "Directly Observing Deuterium" in the [http://www.varianinc.com/image/vimage/docs/products/nmr/apps/pubs/manuals/0199914100a.pdf Deuterium Decoupling Channel Installation] manual with small modifications. To record a 1D deuterium spectrum do the following:
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	##Connect this cable through the BE135-35-8BB filter to the '''XMTR J5313''' port on the broadband preamp		##Connect this cable through the BE135-35-8BB filter to the '''XMTR J5313''' port on the broadband preamp
	#Retrieve parameter set <tt>/vnmr/tests/C13sn</tt>. This will also set <tt>solvent='c6d6'</tt>		#Retrieve parameter set <tt>/vnmr/tests/C13sn</tt>. This will also set <tt>solvent='c6d6'</tt>
	#Make the following changes: <tt>numrfch=4 rftype='dddd' tn='H2' tpwr=43 pw=250 dn=''dm='nnn' dn2='' dm2='nnn' dn3=''dm3='nnn' at=1 d1=1 nt=1 ss=2 in='n'~~''</tt~~</tt>		#Make the following changes: <tt>numrfch=4 rftype='dddd' tn='H2' tpwr=43 pw=250 dn=''dm='nnn' dn2='' dm2='nnn' dn3=''dm3='nnn' at=1 d1=1 nt=1 ss=2 in='n'</tt>
	#<tt>~~Set <tt>~~tof=(7.15-5)*sfrq</tt> to ~~set~~ the carrier closer to the benzene-d6 line		#Set <tt>tof=(7.15-5)*sfrq</tt> to move the carrier closer to the benzene-d6 line
	#Acuire the spectrum with <tt>go</tt> and perform FT with a large <tt>fn</tt> value. You should see a signle line.		#Acuire the spectrum with <tt>go</tt> and perform FT with a large <tt>fn</tt> value. You should see a signle line.
	#Phase the spectrum and center the cursor on the line. Use <tt>movetof</tt> so set the carrier offset on the resonance. Write down the new <tt>tof</tt> value.		#Phase the spectrum and center the cursor on the line. Use <tt>movetof</tt> so set the carrier offset on the resonance. Write down the new <tt>tof</tt> value.
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	~~<br> <br>~~ Even though you can now calibrate the deuterium 90-degree pulse directly with this ~~experment~~, it ~~will~~ not be the one you need. In direct observe mode deuterium is on the first channel, while in a triple-resonance experiment it is on the fourth channel, and the RF path goes through different amplifiers.		Even though you can now calibrate the deuterium 90-degree pulse directly with this experiment, it may not exactly be the one you need for decoupling. In direct observe mode deuterium is on the first channel, while in a triple-resonance experiment it is on the fourth channel, and the RF path goes through different amplifiers.

	=== Indirect Calibration - 1D 13C spectrum ===		=== Indirect Calibration - 1D 13C spectrum ===

	Configure the ~~spectometer~~ for ~~13C~~ direct detection:		Configure the spectrometer for <sup>13</sup>C direct detection:

	On 600 MHz (SUNY Buffalo):		On 600 MHz (SUNY Buffalo):
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	~~<br>~~

	~~==== ====~~

	~~----~~

	==== 13C Anti-Phase Triplet - Indirect Calibration ====		==== 13C Anti-Phase Triplet - Indirect Calibration ====
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	To test 2H decoupling for a protein sample you may take a concentrated DCN sample and run the first increment of HNCA with constant-time Ca evolution, switching deuterium decoupling on and off. If decoupling is working properly, you should see a significant sensitivity difference.		To test 2H decoupling for a protein sample you may take a concentrated DCN sample and run the first increment of HNCA with constant-time Ca evolution, switching deuterium decoupling on and off. If decoupling is working properly, you should see a significant sensitivity difference.

	-~~- Main.AlexEletski - 13 Jan 2007~~		== <span class="mw-headline">'''References''' </span> ==

			<references />

Alex at 22:19, 5 January 2010

2010-01-05T22:19:44Z

← Older revision		Revision as of 22:19, 5 January 2010
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	~~The~~ most ~~common application is~~ deuterium decoupling in triple-resonance experiments with deuterated samples. ~~Any time~~ aliphatic ~~13C~~ magnetization is transverse, ~~you have to decouple~~ it from the attached deuterons. Omitted or improperly calibrated decoupling would cause a sharp sensitivity drop due to scalar relaxation of the second kind (insert reference here).		Pulsing on deuterium is most commonly used as deuterium decoupling in triple-resonance experiments with deuterated samples. Whenever aliphatic <sup>13</sup>C magnetization is transverse, it must be decoupled from the attached deuterons. Omitted or improperly calibrated 2H decoupling would cause a sharp sensitivity drop due to scalar relaxation of the second kind (insert reference here).

	Proper setup requires tuning the 2H channel and calibrating 2H 90-degree pulse.		Proper setup requires tuning the 2H channel and calibrating 2H 90-degree pulse.

Alex at 18:19, 4 January 2010

2010-01-04T18:19:17Z

← Older revision		Revision as of 18:19, 4 January 2010
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	==== 13C Anti-Phase Triplet - Indirect Calibration ====		==== 13C Anti-Phase Triplet - Indirect Calibration ====
			[[Image:antiphase_c6d6.png\|500px\|right]]
			<img alt="antiphase_c6d6.png" src="%ATTACHURL%/antiphase_c6d6.png" />

	Configure the pulse sequence to generate an anti-phase triplet.		Configure the pulse sequence to generate an anti-phase triplet.
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	#Observe the antiphase triplet decrease linearly and then increase inverted.<br>If you do not see any difference between scans you probably do not have pulsing on deuterium.<br>Type <tt>rftype='dddd' BPsetampmode</tt> to fix it and repeat the experiment.		#Observe the antiphase triplet decrease linearly and then increase inverted.<br>If you do not see any difference between scans you probably do not have pulsing on deuterium.<br>Type <tt>rftype='dddd' BPsetampmode</tt> to fix it and repeat the experiment.
	#The zero crossing point corresponds to the desired 90 pulse width. Adjust <tt>dpwr3</tt> so that <tt>pwx3</tt> is ~300 us and repeat the calibration.		#The zero crossing point corresponds to the desired 90 pulse width. Adjust <tt>dpwr3</tt> so that <tt>pwx3</tt> is ~300 us and repeat the calibration.

	~~<img alt="antiphase_c6d6.png" src="%ATTACHURL%/antiphase_c6d6.png" />~~

	Fig. 4. Anti-phase triplet with the middle line suppressed.		Fig. 4. Anti-phase triplet with the middle line suppressed.

Alex at 18:17, 4 January 2010

2010-01-04T18:17:05Z

← Older revision		Revision as of 18:17, 4 January 2010
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	The current protocol is based on the "Directly Observing Deuterium" in the [http://www.varianinc.com/image/vimage/docs/products/nmr/apps/pubs/manuals/0199914100a.pdf Deuterium Decoupling Channel Installation] manual with small modifications. To record a 1D deuterium spectrum do the following:		The current protocol is based on the "Directly Observing Deuterium" in the [http://www.varianinc.com/image/vimage/docs/products/nmr/apps/pubs/manuals/0199914100a.pdf Deuterium Decoupling Channel Installation] manual with small modifications. To record a 1D deuterium spectrum do the following:

			[[Image:C6D6 2H 1D spectrum.png\|right\|500px\|1D deuterium spectrum of C6D6]]

	#Turn lock off and connect the cable from '''J1 PROBE''' port on the 2H decoupler/lock diplexer to the '''J5311 PROBE''' probe port on the broadband preamp with the 2H RF filter in line.		#Turn lock off and connect the cable from '''J1 PROBE''' port on the 2H decoupler/lock diplexer to the '''J5311 PROBE''' probe port on the broadband preamp with the 2H RF filter in line.
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	#Re-connect all cables to their original positions and activate lock		#Re-connect all cables to their original positions and activate lock

	~~<br>~~

	~~[[Image:C6D6 2H 1D spectrum.png\|left\|587x329px\|1D deuterium spectrum of C6D6]]~~

	~~<br>~~

	<br> <br> Even though you can now calibrate the deuterium 90-degree pulse directly with this experment, it will not be the one you need. In direct observe mode deuterium is on the first channel, while in a triple-resonance experiment it is on the fourth channel, and the RF path goes through different amplifiers.		<br> <br> Even though you can now calibrate the deuterium 90-degree pulse directly with this experment, it will not be the one you need. In direct observe mode deuterium is on the first channel, while in a triple-resonance experiment it is on the fourth channel, and the RF path goes through different amplifiers.
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	==== 13C In-Phase Triplet ====		==== 13C In-Phase Triplet ====

			[[Image:Inphase c6d6.png\|500px\|right]]

			[[Image:C6d6 pw90 c13.png\|500px\|right]]


	At this step you will need the <tt>H2_pwxcal</tt> dataset. If you don't have this dataset, install it:		At this step you will need the <tt>H2_pwxcal</tt> dataset. If you don't have this dataset, install it:
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	#Using the current dataset calibrate the 13C 90-degree pulse with the 360 method, along the lines described for 1H (1H cal).<br>Adjust <tt>tpwr</tt> so that <tt>pw</tt> is equal or greater then the typical <tt>pwC</tt> pulse.		#Using the current dataset calibrate the 13C 90-degree pulse with the 360 method, along the lines described for 1H (1H cal).<br>Adjust <tt>tpwr</tt> so that <tt>pw</tt> is equal or greater then the typical <tt>pwC</tt> pulse.

	~~Keep in mind~~ that 13C relaxation is ~~so strong due to 2H quadrupolar mechanism that the effects~~ on the accuracy of pulse width calibration ~~are~~ even stronger than for 1H.		Please note that 13C T1 relaxation is very slow, and its effect on the accuracy of 13C pulse width calibration is even stronger than for 1H.

	~~[[Image:Inphase c6d6.png\|frame\|left]]~~

	~~[[Image:C6d6 pw90 c13.png\|frame\|left]]~~

	<br>		<br>

Aringh at 17:48, 14 December 2009

2009-12-14T17:48:20Z

← Older revision		Revision as of 17:48, 14 December 2009
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	== 2H 90° Automatic Calibration ==		== 2H 90° Automatic Calibration ==

	See ~~the~~ Chapter 9.0 '''2H AutoCalibration''' in the BioPack manual.		See 'Chapter 9.0: '''2H AutoCalibration'''' in the BioPack manual.

	The automatic calibration method of BioPack uses the same protein sample and calibrates the pulse directly on the HDO signal. It requires a K5022 relay to be present, which allows pulsing deuterium on the 4th channel, while detecting it. At SUNY Buffalo 600 MHz and 750 MHz instruments don't have this relay, therefore requiring manual calibration.		The automatic calibration method of BioPack uses the same protein sample and calibrates the pulse directly on the HDO signal. It requires a K5022 relay to be present, which allows pulsing deuterium on the 4th channel, while detecting it. At SUNY Buffalo 600 MHz and 750 MHz instruments don't have this relay, therefore requiring manual calibration.
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	[[Image:C6D6 2H 1D spectrum.png\|left\|587x329px\|1D deuterium spectrum of C6D6]]		[[Image:C6D6 2H 1D spectrum.png\|left\|587x329px\|1D deuterium spectrum of C6D6]]

	<br ~~style="clear: both" /~~>		<br>

	<br> <br> Even though you can now calibrate the deuterium 90-degree pulse directly with this experment, it will not be the one you need. In direct observe mode deuterium is on the first channel, while in a triple-resonance experiment it is on the fourth channel, and the RF path goes through different amplifiers.		<br> <br> Even though you can now calibrate the deuterium 90-degree pulse directly with this experment, it will not be the one you need. In direct observe mode deuterium is on the first channel, while in a triple-resonance experiment it is on the fourth channel, and the RF path goes through different amplifiers.

	=== Indirect Calibration - 1D 13C spectrum ===		=== Indirect Calibration - 1D 13C spectrum ===
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	[[Image:C6d6 pw90 c13.png\|frame\|left]]		[[Image:C6d6 pw90 c13.png\|frame\|left]]

	<br ~~style="clear: both" /~~>		<br>

	==== ====		==== ====
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	Setting '''tof''' to the center line is important here, otherwise the triplet may not be phased properly.		Setting '''tof''' to the center line is important here, otherwise the triplet may not be phased properly.

	Adjust '''pw''' to make the center line disappear.		Adjust '''pw''' to make the center line disappear.

	==== Deuterium Decoupling Test and Probefile Update ====		==== Deuterium Decoupling Test and Probefile Update ====

Alex at 20:28, 11 November 2009

2009-11-11T20:28:25Z

← Older revision		Revision as of 20:28, 11 November 2009
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	Keep in mind that 13C relaxation is so strong due to 2H quadrupolar mechanism that the effects on the accuracy of pulse width calibration are even stronger than for 1H.		Keep in mind that 13C relaxation is so strong due to 2H quadrupolar mechanism that the effects on the accuracy of pulse width calibration are even stronger than for 1H.

	[[Image:Inphase c6d6.png\|frame\|left~~\|13C in-phase triplet of C6D6~~]]		[[Image:Inphase c6d6.png\|frame\|left]]

	[[Image:C6d6 pw90 c13.png\|frame\|left~~\|13C pulse width arrayed from 0 to 100 us. Note the assymmetric profile due to slow 13C relaxation~~]]		[[Image:C6d6 pw90 c13.png\|frame\|left]]

	<br>		<br style="clear: both" />

	==== ====		==== ====

Alex at 19:56, 11 November 2009

2009-11-11T19:56:28Z

← Older revision		Revision as of 19:56, 11 November 2009
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	<br>		<br>

			==== ====

			----

	==== 13C Anti-Phase Triplet - Indirect Calibration ====		==== 13C Anti-Phase Triplet - Indirect Calibration ====
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	Setting '''tof''' to the center line is important here, otherwise the triplet may not be phased properly.		Setting '''tof''' to the center line is important here, otherwise the triplet may not be phased properly.

	Adjust '''pw''' to make the center line disappear.		Adjust '''pw''' to make the center line disappear.

	==== Deuterium Decoupling Test and Probefile Update ====		==== Deuterium Decoupling Test and Probefile Update ====

← Older revision		Revision as of 23:04, 5 January 2010
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	==== 13C Anti-Phase Triplet - Indirect Calibration ====		==== 13C Anti-Phase Triplet - Indirect Calibration ====
	[[Image:antiphase_c6d6.png\|500px\|right]]		[[Image:antiphase_c6d6.png\|500px\|right]]
	~~<img alt="antiphase_c6d6.png" src="%ATTACHURL%/antiphase_c6d6.png" />~~

	Configure the pulse sequence to generate an anti-phase triplet.		Configure the pulse sequence to generate an anti-phase triplet.
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	#Set <tt>dof3</tt> to the <tt>tof</tt> value determined from deuterium 1D spectrum		#Set <tt>dof3</tt> to the <tt>tof</tt> value determined from deuterium 1D spectrum
	#Run experiment with <tt>go</tt>		#Run experiment with <tt>go</tt>
	#Perform FT - you should see the outer lines in anti-phase and the middle line is missing		#Perform FT - you should see the outer lines in anti-phase and the middle line is missing as in the figure on the right.

	The pulse sequence now will applies 90x pulse on 13C followed by a delay tau=1/(4*1J_CD) and another 90x pulse on 13C. The outer lines will evolve during tau to antiphase with respect to 2H. Note that the J-coupling evolution here is "twice as fast" as with two spins 1/2! The second 13C pulse is used to suppress the middle line, which does not evolve under J-coupling.		The pulse sequence now will applies 90x pulse on 13C followed by a delay tau=1/(4*1J_CD) and another 90x pulse on 13C. The outer lines will evolve during tau to antiphase with respect to 2H. Note that the J-coupling evolution here is "twice as fast" as with two spins 1/2! The second 13C pulse is used to suppress the middle line, which does not evolve under J-coupling.


			[[Image:h2_pulse_calib.png\|500px\|right]]
	Now you can calibrate the deuterium pulse width:		Now you can calibrate the deuterium pulse width:

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	#Observe the antiphase triplet decrease linearly and then increase inverted.<br>If you do not see any difference between scans you probably do not have pulsing on deuterium.<br>Type <tt>rftype='dddd' BPsetampmode</tt> to fix it and repeat the experiment.		#Observe the antiphase triplet decrease linearly and then increase inverted.<br>If you do not see any difference between scans you probably do not have pulsing on deuterium.<br>Type <tt>rftype='dddd' BPsetampmode</tt> to fix it and repeat the experiment.
	#The zero crossing point corresponds to the desired 90 pulse width. Adjust <tt>dpwr3</tt> so that <tt>pwx3</tt> is ~300 us and repeat the calibration.		#The zero crossing point corresponds to the desired 90 pulse width. Adjust <tt>dpwr3</tt> so that <tt>pwx3</tt> is ~300 us and repeat the calibration.

	~~Fig. 4. Anti-phase triplet with the middle line suppressed.~~

	~~<img width="834" alt="h2_pulse_calib.png" src="%ATTACHURL%/h2_pulse_calib.png" height="286" />~~

	Fig. 5. Deuterium pulse width calibration. When deuterium pulse with is exactly 90 the outer lines are converted to unobservable MQ coherence.		Fig. 5. Deuterium pulse width calibration. When deuterium pulse with is exactly 90 the outer lines are converted to unobservable MQ coherence.
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	Setting '''tof''' to the center line is important here, otherwise the triplet may not be phased properly.		Setting '''tof''' to the center line is important here, otherwise the triplet may not be phased properly.

	Adjust '''pw''' to make the center line disappear.		Adjust '''pw''' to make the center line disappear.

	==== Deuterium Decoupling Test and Probefile Update ====		==== Deuterium Decoupling Test and Probefile Update ====