Print all pages in /TPAD2.0/Project 1/Test Plate Experiment

TEST PLATE	1	2	3	4	5	6	7	8	9	10	11	12
A	Batch Ref (VP)	141171	141201	613	151280	141361	1231	1095	141374	Batch QC (GW)	141345	1091
B	Batch QC (GW)	1207	141232	587	141335	1116	Batch Ref (VP)	1030	1197	151068	1014	141112
C	687	141318	Batch QC (GW)	987	141229	1205	151009	141352	141364	629	Batch Ref (VP)	481
D	1211	151217	131276	141202	151178	Batch QC (GW)	592	Batch Ref (VP)	141346	1226	151019	301
E	1175	1111	1236	Batch Ref (VP)	534	141130	Batch QC (GW)	141091	141348	141337	141362	141372
F	141365	Batch Ref (VP)	151187	1262	141440	141236	1189	1034	Batch QC (GW)	638	141088	1132
G	532	1210	141387	141078	1171	Batch Ref (VP)	141217	Batch QC (GW)	1018	1163	1237	141370
H	1133	151034	141099	151190	Batch QC (GW)	1167	141353	1113	1078	141070	1229	Batch Ref (VP)

	n
Batch Ref	8	Ventricular Pool
Healthy Control	51	Group 1
AD/MCI	14	Group 2
PD/PDI-MCI/LBD	15	Group 3
Batch QC	8	Golden West

Row 1		Row 2		Row 3		Row 4		Row 5		Row 6		Row 7		Row 8
Batch Ref (VP)	B1VP-01	Batch QC (GW)	B1QC-02	687	ADC1-21	1211	ADC1-31	1175	ADC1-41	141365	ADC1-51	532	ADC1-61	1133	ADC1-71
141171	ADC1-01	1207	ADC1-11	141318	ADC1-22	151217	ADC1-32	1111	ADC1-42	Batch Ref (VP)	B1VP-06	1210	ADC1-62	151034	ADC1-72
141201	ADC1-02	141232	ADC1-12	Batch QC (GW)	B1QC-03	131276	ADC1-33	1236	ADC1-43	151187	ADC1-52	141387	ADC1-63	141099	ADC1-73
613	ADC1-03	587	ADC1-13	987	ADC1-23	141202	ADC1-34	Batch Ref (VP)	B1VP-05	1262	ADC1-53	141078	ADC1-64	151190	ADC1-74
151280	ADC1-04	141335	ADC1-14	141229	ADC1-24	151178	ADC1-35	534	ADC1-44	141440	ADC1-54	1171	ADC1-65	Batch QC (GW)	B1QC-08
141361	ADC1-05	1116	ADC1-15	1205	ADC1-25	Batch QC (GW)	B1QC-04	141130	ADC1-45	141236	ADC1-55	Batch Ref (VP)	B1VP-07	1167	ADC1-75
1231	ADC1-06	Batch Ref (VP)	B1VP-02	151009	ADC1-26	592	ADC1-36	Batch QC (GW)	B1QC-05	1189	ADC1-56	141217	ADC1-66	141353	ADC1-76
1095	ADC1-07	1030	ADC1-16	141352	ADC1-27	Batch Ref (VP)	B1VP-04	141091	ADC1-46	1034	ADC1-57	Batch QC (GW)	B1QC-07	1113	ADC1-77
141374	ADC1-08	1197	ADC1-17	141364	ADC1-28	141346	ADC1-37	141348	ADC1-47	Batch QC (GW)	B1QC-06	1018	ADC1-67	1078	ADC1-78
Batch QC (GW)	B1QC-01	151068	ADC1-18	629	ADC1-29	1226	ADC1-38	141337	ADC1-48	638	ADC1-58	1163	ADC1-68	141070	ADC1-79
141345	ADC1-09	1014	ADC1-19	Batch Ref (VP)	B1VP-03	151019	ADC1-39	141362	ADC1-49	141088	ADC1-59	1237	ADC1-69	1229	ADC1-80
1091	ADC1-10	141112	ADC1-20	481	ADC1-30	301	ADC1-40	141372	ADC1-50	1132	ADC1-60	141370	ADC1-70	Batch Ref (VP)	B1VP-08

B1VP-01	ADC1-01	ADC1-02	ADC1-03	ADC1-04	ADC1-05	ADC1-06	ADC1-07	ADC1-08	B1QC-01	ADC1-09	ADC1-10
B1QC-02	ADC1-11	ADC1-12	ADC1-13	ADC1-14	ADC1-15	B1VP-02	ADC1-16	ADC1-17	ADC1-18	ADC1-19	ADC1-20
ADC1-21	ADC1-22	B1QC-03	ADC1-23	ADC1-24	ADC1-25	ADC1-26	ADC1-27	ADC1-28	ADC1-29	B1VP-03	ADC1-30
ADC1-31	ADC1-32	ADC1-33	ADC1-34	ADC1-35	B1QC-04	ADC1-36	B1VP-04	ADC1-37	ADC1-38	ADC1-39	ADC1-40
ADC1-41	ADC1-42	ADC1-43	B1VP-05	ADC1-44	ADC1-45	B1QC-05	ADC1-46	ADC1-47	ADC1-48	ADC1-49	ADC1-50
ADC1-51	B1VP-06	ADC1-52	ADC1-53	ADC1-54	ADC1-55	ADC1-56	ADC1-57	B1QC-06	ADC1-58	ADC1-59	ADC1-60
ADC1-61	ADC1-62	ADC1-63	ADC1-64	ADC1-65	B1VP-07	ADC1-66	B1QC-07	ADC1-67	ADC1-68	ADC1-69	ADC1-70
ADC1-71	ADC1-72	ADC1-73	ADC1-74	B1QC-08	ADC1-75	ADC1-76	ADC1-77	ADC1-78	ADC1-79	ADC1-80	B1VP-08

MAKE SURE HEATING PLATES/BLOCKS ARE AT CORRECT TEMP

MATERIALS:

Pooled ventricular CSF; 8 60 ul aliquots
GW CSF; 3 250 ul aliquots
BCA reagent; 22,000 ul total

21,560 ul of reagent A
440 ul of reagent B

1M iodoacetamide (IAA) (1 plate)
1M DL-dithiothreitol (DTT) (1 plate)
50 mM Tris, pH 8.5

30 mL

1.5 mL 50 1M Tris + 28.5 mL water; pH to 8.5

2% SDS

Dilute 20% SDS with Tris pH 8.5
2,000 ul 20% SDS in 18,000 ul Tris

Resyn Biosciences MagResyn Hydroxyl particles
100% Acetonitrile (ACN)
70% Ethanol

200 mL

95% Acetonitrile

300 mL

100 ng/ul enolase

1,750 ul (0.2 ug/ul) aliquot in freezer 3
Thaw, add 1,600 ul of it to reservoir with 1,600 ul of lysis buffer

50 mM ammonium bicarbonate (ABC)
Pierce Porcine trypsin; 3 100-ug vials

Add 2,000 ul of 50 mM ABC to each vial to get working concentration of 0.05 ug/ul

HPLC grade 100% formic acid
500 fmol Pierce Retention Time Calibrant

Dilute 5 pmol PRTC (stock) to 500 fmol

Materials cont'd

Labeled binding, wash, and elution plates
Labeled reservoirs

METHODS:

Protein Assay

Thaw everything, and put on ice in order in 4 buckets

Perform BCA assay on the to determine protein concentration

200 ul of BCA reagent into each well of the assay plate

*3 rows per plate, then bring it to the 37 degree room, so 4 total plates

14 of 2% SDS into the 4 sample wells

Add 2 ul of sample to the wells

Add 16 ul of each standard to the wells

Pipette up and down to mix and heat for 30 min at 37 C

Use Varioskan flash to obtain photometric reading of the plates

Calculations

Whichever sample has the lowest concentration will be the baseline, so all other samples will be brought up to this volume with DPBS

So the amount of 2% SDS added to every sample will be the same

sample	conc	amt need for 25 ug	amt DPBS needed to bring to lowest conc vol	ul enolase	vol 2% SDS to add	1M IAA	1M DTT	vol for 70% final conc of ACN
Batch Ref (VP)	.896	27.9	6.39	8	34.29	3.06	1.6
141171	.941	26.56	7.73	8	34.29	3.06	1.6
141201	1.204	20.77	13.52	8	34.29	3.06	1.6
613	.769	32.5	1.79	8	34.29	3.06	1.6
151280	.769	32.5	1.79	8	34.29	3.06	1.6
141361	.914	27.35	6.94	8	34.29	3.06	1.6
1231	.91	27.49	6.8	8	34.29	3.06	1.6
1095	.846	29.55	4.74	8	34.29	3.06	1.6
141374	.819	30.52	3.77	8	34.29	3.06	1.6
Batch QC (GW)	.95	26.31	7.98	8	34.29	3.06	1.6
141345	.973	25.7	8.59	8	34.29	3.06	1.6
1091	.878	28.48	5.81	8	34.29	3.06	1.6
Batch QC (GW)	.86	29.08	5.21	8	34.29	3.06	1.6
1207	.882	28.33	5.96	8	34.29	3.06	1.6
141232	.887	28.19	6.1	8	34.29	3.06	1.6
587	.869	28.78	5.51	8	34.29	3.06	1.6
141335	.742	33.69	.6	8	34.29	3.06	1.6
1116	1.122	22.28	12.01	8	34.29	3.06	1.6
Batch Ref (VP)	.946	26.44	7.85	8	34.29	3.06	1.6
1030	.882	28.33	5.96	8	34.29	3.06	1.6
1197	1.	25.	9.29	8	34.29	3.06	1.6
151068	.937	26.69	7.6	8	34.29	3.06	1.6
1014	.914	27.35	6.94	8	34.29	3.06	1.6
141112	.869	28.78	5.51	8	34.29	3.06	1.6
687	8.416	2.97	31.32	8	34.29	3.06	1.6
141318	.869	28.78	5.51	8	34.29	3.06	1.6
Batch QC (GW)	1.054	23.71	10.58	8	34.29	3.06	1.6
987	.991	25.23	9.06	8	34.29	3.06	1.6
141229	.882	28.33	5.96	8	34.29	3.06	1.6
1205	.864	28.93	5.36	8	34.29	3.06	1.6
151009	.878	28.48	5.81	8	34.29	3.06	1.6
141352	.742	33.69	.6	8	34.29	3.06	1.6
141364	.824	30.36	3.93	8	34.29	3.06	1.6
629	.91	27.49	6.8	8	34.29	3.06	1.6
Batch Ref (VP)	.919	27.22	7.07	8	34.29	3.06	1.6
481	.792	31.57	2.72	8	34.29	3.06	1.6
1211	.946	26.42	7.87	8	34.29	3.06	1.6
151217	.907	27.57	6.72	8	34.29	3.06	1.6
131276	1.132	22.08	12.21	8	34.29	3.06	1.6
141202	.828	30.18	4.11	8	34.29	3.06	1.6
151178	.824	30.36	3.93	8	34.29	3.06	1.6
Batch QC (GW)	.892	28.02	6.27	8	34.29	3.06	1.6
592	.926	26.98	7.31	8	34.29	3.06	1.6
Batch Ref (VP)	.941	26.56	7.73	8	34.29	3.06	1.6
141346	.961	26.02	8.27	8	34.29	3.06	1.6
1226	.804	31.1	3.19	8	34.29	3.06	1.6
151019	.892	28.02	6.27	8	34.29	3.06	1.6
301	1.015	24.64	9.65	8	34.29	3.06	1.6
1175	.877	28.49	5.8	8	34.29	3.06	1.6
1111	.819	30.54	3.75	8	34.29	3.06	1.6
1236	.838	29.82	4.47	8	34.29	3.06	1.6
Batch Ref (VP)	.985	25.37	8.92	8	34.29	3.06	1.6
534	.755	33.12	1.17	8	34.29	3.06	1.6
141130	1.01	24.76	9.53	8	34.29	3.06	1.6
Batch QC (GW)	.877	28.49	5.8	8	34.29	3.06	1.6
141091	.819	30.54	3.75	8	34.29	3.06	1.6
141348	.838	29.82	4.47	8	34.29	3.06	1.6
141337	.863	28.98	5.31	8	34.29	3.06	1.6
141362	1.064	23.5	10.79	8	34.29	3.06	1.6
141372	.961	26.02	8.27	8	34.29	3.06	1.6
141365	.941	26.56	7.73	8	34.29	3.06	1.6
Batch Ref (VP)	1.157	21.61	12.68	8	34.29	3.06	1.6
151187	.907	27.57	6.72	8	34.29	3.06	1.6
1262	.843	29.65	4.64	8	34.29	3.06	1.6
141440	.99	25.25	9.04	8	34.29	3.06	1.6
141236	.917	27.27	7.02	8	34.29	3.06	1.6
1189	.799	31.29	3.	8	34.29	3.06	1.6
1034	.833	30.	4.29	8	34.29	3.06	1.6
Batch QC (GW)	.912	27.42	6.87	8	34.29	3.06	1.6
638	.892	28.02	6.27	8	34.29	3.06	1.6
141088	.912	27.42	6.87	8	34.29	3.06	1.6
1132	.961	26.02	8.27	8	34.29	3.06	1.6
532	.802	31.17	3.12	8	34.29	3.06	1.6
1210	.729	34.29	.	8	34.29	3.06	1.6
141387	.844	29.63	4.66	8	34.29	3.06	1.6
141078	.875	28.57	5.72	8	34.29	3.06	1.6
1171	.823	30.38	3.91	8	34.29	3.06	1.6
Batch Ref (VP)	.875	28.57	5.72	8	34.29	3.06	1.6
141217	.943	26.52	7.77	8	34.29	3.06	1.6
Batch QC (GW)	.88	28.4	5.89	8	34.29	3.06	1.6
1018	.786	31.79	2.5	8	34.29	3.06	1.6
1163	.828	30.19	4.1	8	34.29	3.06	1.6
1237	.974	25.67	8.62	8	34.29	3.06	1.6
141370	.891	28.07	6.22	8	34.29	3.06	1.6
1133	.833	30.	4.29	8	34.29	3.06	1.6
151034	.932	26.82	7.47	8	34.29	3.06	1.6
141099	1.	25.	9.29	8	34.29	3.06	1.6
151190	1.208	20.69	13.6	8	34.29	3.06	1.6
Batch QC (GW)	1.	25.	9.29	8	34.29	3.06	1.6
1167	1.208	20.69	13.6	8	34.29	3.06	1.6
141353	.953	26.23	8.06	8	34.29	3.06	1.6
1113	.969	25.81	8.48	8	34.29	3.06	1.6
1078	.901	27.75	6.54	8	34.29	3.06	1.6
141070	.938	26.67	7.62	8	34.29	3.06	1.6
1229	1.083	23.08	11.21	8	34.29	3.06	1.6
Batch Ref (VP)	1.	25.	9.29	8	34.29	3.06	1.6

Protein denaturation

To each well add a volume of 2% SDS equivalent to the volume of CSF added

Pipette up and down gently

Add DPBS to bring up samples to the volume equivalent to the most dilute sample

Sample preparation

Based on the concentrations, pull 25 ug and add to plate

Add 8 ul of 100 ng/ul enolase

Reduction and alkylation

Volumes of reducing and alkylating agents will be calculated after the sample and buffer volumes are determined (above)

Add ____ ul of 1M DTT to each well, pipette up and down gently

Heat the plate at 95C for 10 min

Allow to cool

Add ____ ul of 1M IAA to each well

Pipette up and down gently

Incubate at room temperature in the dark for 30 minutes

Protein precipitation

Add 12.5 ul of the Hydroxyl beads (at their stock concentration of 20 ug/ul) to each well

Add _____ ul of 100% ACN to each well (70% final concentration) to precipitate proteins

Allow to sit for 10 minutes at room temperature

NOTE: You cannot freeze the samples at this point--the hydroxyl beads will rupture when frozen.

Wash and digestion plate preparation

Wash plates (deep well):

Washes 1-3; 1 mL 95% ACN
Washes 4-5; 1 mL EtOH

Digestion/elution plate (deep well):

100 ul 50 mM ABC + 50 ul of 0.05 ug/ul Pierce Porcine trypsin (1:10 enzyme:protein ratio)

Kingfisher automated clean-up and digestion

Turn on Kingfisher and bring up adapted Resyn protocol (see attached)

Make sure binding plate volume has been adjusted

Add plates to instrument as indicated by the program

Hit "run"

The automated clean-up and digestion will take about 90 minutes.

Quenching the digestion

Add 5.4 ul of 100% formic acid to quench

Freeze (-80C) or dry down in speed vacuum

In my case, I transferred the samples from the 96 well plate and into 8 individual 1.5 mL tubes (NOT lo-bind)

By clicking through these pages you can find things such as protocol, run logs, meta data, etc

Pages

Replicate	Linkcode	Diagnosis	Group	Age	isMale	Cutpoint	ptau181 (pg/ml)	Ttau (pg/ml)	AB42/AB40	AB42 (pg/ml)	Abeta40	Y1 Consensus	Y2 Consensus	Y3 Consensus	Y4 Consensus	PlateRow
ADC1-01	141171	AD	2	68	0	Cutpoint #1	70.3	532.3	0.09	613.3	7185					1
ADC1-02	141201	PD-MCI	3	85.8	1	Cutpoint #2	158	993	0.06	879	15201					1
ADC1-03	613	HC	1	67	1	Cutpoint #1	26.2	206.8	0.11	1131.3	9856.9	HC	HC	HC	HC	1
ADC1-04	151280	HC	1	79	0	Cutpoint #1	49.5	364.8	0.11	1346.3	12418.9					1
ADC1-05	141361	HC	1	65.2	0	Cutpoint #2	30.6	196	0.12	961	7836					1
ADC1-06	1231	HC	1	67	0	Cutpoint #1	59	409.8	0.13	1598.3	12370.9					1
ADC1-07	1095	HC	1	67	1	Cutpoint #1	37.7	267.8	0.09	977.3	10715.9					1
ADC1-08	141374	PD-noMCI	3	73.3	0	Cutpoint #2	33.8	207	0.09	899	9769					1
ADC1-09	141345	HC	1	79.3	0	Cutpoint #2	27.4	169	0.13	806	6188					1
ADC1-10	1091	MCI	2	78	1	Cutpoint #1	54	367	0.06	460	7580	MCI	MCI			1
ADC1-11	1207	HC	1	78	1	Cutpoint #1	67.1	423	0.08	1000	12425	HC	HC			2
ADC1-12	141232	HC	1	70.6	0	Cutpoint #2	48.6	294	0.06	613	9780					2
ADC1-13	587	MCI	2	78	1	Cutpoint #1	152.7	844	0.06	918	16016	MCI	MCI			2
ADC1-14	141335	PDD	3	64.7	1	Cutpoint #2	31.2	178	0.08	345	4191					2
ADC1-15	1116	HC	1	70	1	Cutpoint #1	43.7	295.8	0.12	1206.3	10045.9					2
ADC1-16	1030	HC	1	66	0	Cutpoint #1	63.2	478.8	0.13	1941.3	15198.9					2
ADC1-17	1197	MCI	2	79	1	Cutpoint #1	23.1	362	0.13	1047	8214	MCI				2
ADC1-18	151068	HC	1	84	0	Cutpoint #1	37.2	277.8	0.12	997.3	8245.9					2
ADC1-19	1014	HC	1	68	0	Cutpoint #1	47.3	361.8	0.12	1707.3	14269.9					2
ADC1-20	141112	PD-MCI	3	65.8	1	Cutpoint #2	16.4	93	0.11	564	4948					2
ADC1-21	687	HC	1	76	0	Cutpoint #1	38.5	289.8	0.13	1532.3	11787.9	HC	HC	HC	HC	3
ADC1-22	141318	PD-MCI	3	76	1	Cutpoint #2	55.7	345	0.06	383	6092					3
ADC1-23	987	HC	1	72	1	Cutpoint #1	33.7	259.8	0.11	1082.3	9706.9					3
ADC1-24	141229	HC	1	59.7	1	Cutpoint #2	26.8	181	0.13	921	7113					3
ADC1-25	1205	AD	2	63	0	Cutpoint #1	147.4	882	0.07	898	12586	AD	AD			3
ADC1-26	151009	HC	1	73	1	Cutpoint #1	44.5	316.8	0.09	982.3	11149.9					3
ADC1-27	141352	PD-noMCI	3	58.4	0	Cutpoint #2	26.8	178	0.1	592	5958					3
ADC1-28	141364	HC	1	74	0	Cutpoint #1	47.6	360.8	0.12	1259.3	10791.9					3
ADC1-29	629	HC	1	79	1	Cutpoint #1	34.9	244.8	0.12	790.3	6728.9	HC	HC	HC	HC	3
ADC1-30	481	HC	1	75	0	Cutpoint #1	33.8	243.8	0.12	1056.3	9022.9					3
ADC1-31	1211	MCI	2	74	0	Cutpoint #1	85.3	533	0.06	647	10629	MCI	MCI			4
ADC1-32	151217	HC	1	74	0	Cutpoint #1	37.9	357.8	0.11	1111.3	9776.9					4
ADC1-33	131276	HC	1	81	1	Cutpoint #1	45.5	323.8	0.1	1175.3	11316.9					4
ADC1-34	141202	PDD	3	69.3	0	Cutpoint #2	80.1	525	0.08	503	6262					4
ADC1-35	151178	HC	1	73	1	Cutpoint #1	31.2	275.8	0.13	790.3	6257.9					4
ADC1-36	592	HC	1	81	1	Cutpoint #1	35.9	259.8	0.09	790.3	8659.9					4
ADC1-37	141346	HC	1	64.8	1	Cutpoint #2	46.7	343	0.15	1658	11275					4
ADC1-38	1226	MCI	2	70	1	Cutpoint #1	83.5	450	0.07	825	12000	MCI				4
ADC1-39	151019	PDD	3	75.2	1	Cutpoint #2	21.2	131	0.09	326	3717					4
ADC1-40	301	HC	1	71	1	Cutpoint #1	43.5	371.8	0.12	1560.3	12734.9					4
ADC1-41	1175	HC	1	70	1	Cutpoint #1	40.1	1325	0.09	932	10654	HC	HC			5
ADC1-42	1111	HC	1	70	0	Cutpoint #1	26.2	217	0.13	1143	9078	HC	HC			5
ADC1-43	1236	AD	2	82	0	Cutpoint #1	113.9	776.3	0.07	638.3	9806	AD	AD			5
ADC1-44	534	HC	1	70	0	Cutpoint #1	17.7	132.8	0.15	685.3	4572.9					5
ADC1-45	141130	PD-noMCI	3	73.5	1	Cutpoint #2	26.4	165	0.14	1072	7827					5
ADC1-46	141091	HC	1	75	1	Cutpoint #2	30.2	178	0.12	959	7815					5
ADC1-47	141348	MCI	2	75	0	Cutpoint #1	139.2	975.8	0.05	864.3	16860.9					5
ADC1-48	141337	HC	1	69.5	0	Cutpoint #2	52.6	359	0.12	1478	11910					5
ADC1-49	141362	HC	1	62	0	Cutpoint #2	36.5	269	0.13	1292	9874					5
ADC1-50	141372	PD-noMCI	3	62.5	0	Cutpoint #2	19	100	0.12	526	4300					5
ADC1-51	141365	PDD	3	72.5	1	Cutpoint #2	35.8	207	0.07	296	4392					6
ADC1-52	151187	HC	1	74	0	Cutpoint #1	52.9	439.8	0.06	643.3	11359.9					6
ADC1-53	1262	AD	2	58	0	Cutpoint #1	127.5	768.3	0.07	639.3	9641	AD				6
ADC1-54	141440	HC	1	83	0	Cutpoint #1	53.2	400.8	0.11	1606.3	14998.9					6
ADC1-55	141236	HC	1	62.3	1	Cutpoint #2	43.1	310	0.12	1476	11873					6
ADC1-56	1189	HC	1	72	0	Cutpoint #1	44	276	0.06	528	8396	HC	HC			6
ADC1-57	1034	AD	2	60	0	Cutpoint #1	129.8	988	0.05	412	7743	AD	AD			6
ADC1-58	638	HC	1	62	0	Cutpoint #1	30.2	214.8	0.12	945.3	7933.9	HC	HC	HC	HC	6
ADC1-59	141088	PD-MCI	3	68	0	Cutpoint #2	26.1	236	0.15	876	6017					6
ADC1-60	1132	HC	1	73	0	Cutpoint #1	35.2	231	0.1	892	9188	HC	HC			6
ADC1-61	532	HC	1	74	0	Cutpoint #1	37.3	306.8	0.11	1041.3	9638.9					7
ADC1-62	1210	HC	1	65	0	Cutpoint #1	12.7	219	0.13	546	4339	HC	HC			7
ADC1-63	141387	HC	1	65.6	0	Cutpoint #2	23.5	169	0.14	909	6731					7
ADC1-64	141078	HC	1	65.9	0	Cutpoint #2	18.8	114	0.13	525	4089					7
ADC1-65	1171	MCI	2	78	1	Cutpoint #1	28.6	219	0.14	1476	10471	MCI	MCI			7
ADC1-66	141217	PD-MCI	3	71	1	Cutpoint #2	36.6	233	0.13	1099	8432					7
ADC1-67	1018	HC	1	68	0	Cutpoint #1	30.7	269.8	0.11	898.3	7842.9					7
ADC1-68	1163	HC	1	68	0	Cutpoint #1	75.7	409.8	0.07	472.3	6468.9					7
ADC1-69	1237	AD	2	73	1	Cutpoint #1	397.9	1926.3	0.04	624.3	14096	AD	AD			7
ADC1-70	141370	HC	1	64.9	0	Cutpoint #2	49.8	373	0.12	1619	12954					7
ADC1-71	1133	HC	1	51	0	Cutpoint #1	34.4	217	0.13	1384	10570	HC				8
ADC1-72	151034	HC	1	83	1	Cutpoint #1	20.1	149.8	0.15	594.3	3973.9					8
ADC1-73	141099	PD-noMCI	3	67.4	1	Cutpoint #2	62.3	363	0.05	603	11146					8
ADC1-74	151190	HC	1	84	0	Cutpoint #1	44.4	358.8	0.12	1408.3	11561.9					8
ADC1-75	1167	HC	1	67	0	Cutpoint #1	35.4	298	0.14	1778	12383	HC	HC			8
ADC1-76	141353	HC	1	81	0	Cutpoint #1	49.4	360.8	0.1	1169.3	11338.9					8
ADC1-77	1113	HC	1	73	1	Cutpoint #1	33.1	229	0.14	1155	8167	HC	HC			8
ADC1-78	1078	MCI	2	83	0	Cutpoint #1	67.8	456	0.07	926	13142	MCI	MCI			8
ADC1-79	141070	PD-MCI	3	73.7	1	Cutpoint #2	48.3	325	0.08	490	6139					8
ADC1-80	1229	HC	1	69	0	Cutpoint #1	48.4	392	0.13	1596	12716	HC	HC			8
B1VP-01	Batch Ref (VP)															1
B1VP-02	Batch Ref (VP)															2
B1VP-03	Batch Ref (VP)															3
B1VP-04	Batch Ref (VP)															4
B1VP-05	Batch Ref (VP)															5
B1VP-06	Batch Ref (VP)															6
B1VP-07	Batch Ref (VP)															7
B1VP-08	Batch Ref (VP)															8
B1QC-01	Batch QC (GW)															1
B1QC-02	Batch QC (GW)															2
B1QC-03	Batch QC (GW)															3
B1QC-04	Batch QC (GW)															4
B1QC-05	Batch QC (GW)															5
B1QC-06	Batch QC (GW)															6
B1QC-07	Batch QC (GW)															7
B1QC-08	Batch QC (GW)															8

The samples were prepped as EvoSep tips in 4 sets, general tip prep protocol is attached. The attached .csv's contain the run order, methods, output directories, etc.

Test plate analysis on Thermo Orbitrap Exploris instrument issue log:

Set 1:

18 May 2021 - 20 May 2021
no failures

Set 2:

20 May 2021 - 21 May 2021
no failures

Set 3:

21 May 2021 - 22 May 2021
no failures

Set 4:

22 May 2021 @ 11:48 PM, over-pressurization failure

Fixed + restarted @ 12:18 PM on 23 May 2021

23 May 2021 @ 11:15 PM overpressurization failure
24 May 2021 @ 1:15 AM power failure

Fixed (needle tip cleaned) + restarted @ 12:52 PM on 24 May 2021

Date of sample prep: 26 April 2021

Samples: 80 CSF samples from the clinical core + 8 batch references + 8 batch QCs

Sample prep protocol: SP3

Instrument: Thermo Exploris

Dates of data collection: 18 May 2021 - 25 May 2021

General description: this test plate is the first batch of human cerebral spinal fluid samples for proteomics as part of the TPAD2.0 project. 96 samples (including 80 experimental samples, 8 batch references and 8 batch QCs) were prepped using SP3 and then analyzed together in a randomized order with 2 sets of chromatogram library injections. With this dataset we aim to confirm the reproducibility of the assay and protocol we developed in Year 1, before embarking on prepping the remaining samples in the project.

Table of Contents

Protocol

Index

Pages

Protocol

Meta data

Run order and notes

Meta data

Run order and notes

Experiment description