Scheme 1B - 2018

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UK NEQAS FOR H&I SCHEME 1B - HLA-B27 TESTING HLA-B27 TYPING RESULTS OF SAMPLES 1B01&02/2018 DESPATCHED ON 06TH MARCH 2018 Summary of Results Total tested (excl. Equivocal)

124

116

Donor HLA type

1B01/2018

HLA-A2, A-; B27, B47; Cw1, Cw6; DR103, DR7, DQ2, DQ7

121 3 3 97.6% 2.4%

111 5 11 95.7% 4.3%

Donor HLA type

1B02/2018

HLA-A3, A-; B27, B60; Cw1, Cw10; DR8, DR11, DQ4, DQ7

Positive

Positive

1 3 4 7

Result 1B01/2018 Positive NT Positive Equivocal

Result 1B02/2018 Positive NT Positive Equivocal

Date received 07-Mar NT 07-Mar 09-Mar

Date tested 16-Mar NT 08-Mar 14-Mar

8 9 10 11 12 14 15 17 19 20 21 22 23 24 25 26 28 29 31 32 34 35 37

Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive

Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive

08-Mar 07-Mar 07-Mar 07-Mar 08-Mar 07-Mar 07-Mar 08-Mar 07-Mar 07-Mar 07-Mar 08-Mar 07-Mar 09-Mar 12-Mar 07-Mar 07-Mar 08-Mar 07-Mar 06-Mar 07-Mar 07-Mar 09-Mar

09-Mar 07-Mar 29-Mar 16-Mar 20-Mar 09-Mar 19-Mar 08-Mar 09-Mar 08-Mar 09-Mar 08-Mar 14-Mar 13-Mar 08-Mar 09-Mar 12-Mar 13-Mar 08-Mar 08-Mar 08-Mar 15-Mar

Serological Molecular Serological Molecular Molecular Molecular Molecular Serological & Molecular Molecular Serological Serological Molecular Molecular Molecular Molecular Molecular Molecular Molecular Serological Serological Molecular Molecular Molecular

39 40 41 42

Positive Positive Positive Positive

Positive Positive Positive Positive

07-Mar 07-Mar 07-Mar 09-Mar

07-Mar 08-Mar 14-Mar 13-Mar

Positive Negative NT/Equivocal % Positive % Negative Consensus

Lab No

Method Primers and Probes

PCR-SSP Detection system

Comments

High Resolution Typing HLA-B*27 alleles 1B01/2018

1B02/2018

Molecular Poor sample quality Serological

Both samples fall within our "equivocal" range. Patient samples would have been sent for confirmation by PCR.

Own design Own design Own design

Gel Gel Gel

Commercial

Fluorescence

Own design Commercial

Gel Gel

Commercial

Gel, BAG reverse dot blot

Commercial

SSOP

Molecular Serological Molecular Molecular

Own design

Gel

Own design

Gel

Own design

45 46

Positive Positive

Positive Equivocal

07-Mar 05-Mar

12-Mar 03-Sep

Molecular Serological

47 48 54 55 57 58 62 65

Positive Positive Positive Positive Positive Positive Positive Equivocal

Positive Positive Positive Positive Equivocal Positive Positive Equivocal

08-Mar 08-Mar 07-Mar 03-Mar 07-Mar 07-Mar 08-Mar 13-Mar

08-Mar 12-Mar 09-Mar 20-Mar 07-Mar 08-Mar 08-Mar 15-Mar

Serological Molecular Molecular Molecular Serological Molecular Molecular Serological

66 67 72 76

Positive Positive Positive Positive

Positive Negative Positive Equivocal

09-Mar 06-Mar 12-Mar 07-Mar

13-Mar 07-Mar 16-Mar 09-Mar

Molecular Serological Molecular Serological

77 78 83 84

Positive Positive Positive Positive

Positive Positive Negative Equivocal

08-Mar 08-Mar 07-Mar 08-Mar

08-Mar 14-Mar 07-Mar 09-Mar

Serological Molecular Serological Serological & Molecular

85 86 90 96 101 103 104

Positive Positive Positive Positive Positive Positive Positive

Positive Positive Positive Positive Positive Positive Equivocal

07-Mar 07-Mar 12-Mar 08-Mar 07-Mar 07-Mar 08-Mar

09-Mar 12-Mar 13-Mar 16-Mar 08-Mar 08-Mar 08-Mar

Molecular Molecular Serological Molecular Molecular Serological Serological

106 107

Negative Positive

Negative Positive

15-Mar 12-Mar

16-Mar 14-Mar

Molecular

108 109 110 111 112 113 115 126 127 128 129 130 133 137 142 150 153 154 156 159 174 176 185

Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive

Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive

12-Mar 07-Mar 07-Mar 07-Mar 12-Mar 07-Mar 08-Mar 12-Mar 08-Mar 07-Mar 08-Mar 09-Mar 09-Mar 07-Mar 07-Mar 06-Mar 07-Mar 07-Mar 08-Mar 08-Mar 19-Mar 12-Mar 07-Mar

15-Mar 09-Mar 07-Mar 08-Mar 16-Mar 12-Mar 12-Mar 19-Mar 21-Mar 22-Mar 09-Mar 09-Mar 12-Mar 07-Mar 13-Mar 08-Mar 08-Mar 09-Mar 12-Mar 16-Mar 21-Mar 12-Mar 13-Mar

Molecular Molecular Serological & Molecular Serological Molecular Molecular Molecular Molecular Molecular Serological & Molecular Molecular Serological & Molecular Molecular Serological Molecular Molecular Serological Molecular Molecular Molecular Molecular Serological Molecular

198 209 219 223 224

Positive Positive Positive Positive Positive

Positive Positive Positive Positive Positive

07-Mar 07-Mar 14-Mar 12-Mar 07-Mar

08-Mar 08-Mar 16-Mar 14-Mar 14-Mar

Serological Molecular

225 232 234 236 245 252 255 256 257 269 273 276 278

Positive Positive Positive Positive Positive Positive Positive Negative Positive Positive Positive Positive Positive

Positive Positive Positive Positive Positive Positive Positive Negative Positive Positive Positive Positive Positive

12-Mar 10-Mar 12-Mar 14-Mar 07-Mar 05-Mar 08-Mar 08-Mar 07-Mar 14-Mar 07-Mar 07-Mar 12-Mar

13-Mar 13-Mar 13-Mar 19-Mar 12-Mar 19-Mar 16-Mar 09-Mar 14-Mar 15-Mar 09-Mar 19-Mar 19-Mar

279 288 292 295 300 305 306 307 315 317 319 324 327 334 339 340 348

Negative Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive

Negative Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Positive Equivocal

07-Mar 07-Mar 12-Mar 07-Mar 16-Mar 14-Mar 08-Mar 07-Mar 08-Mar 12-Mar 07-Mar 14-Mar 07-Mar 08-Mar 07-Mar 08-Mar 12-Mar

09-Mar 07-Mar 13-Mar 07-Mar 17-Mar 16-Mar 15-Mar 12-Mar 14-Mar 15-Mar 08-Mar 14-Mar 19-Mar 08-Mar 12-Mar 08-Mar 13-Mar

Serological Serological Serological Serological Molecular

349 353 357

Positive Positive Positive

Positive Positive Equivocal

08-Mar 08-Mar 09-Mar

08-Mar 19-Mar 09-Mar

Serological Molecular Serological

359 362 365 368 372 373 377 378

Positive Positive Positive Positive Positive Positive Positive Positive

Positive Positive Equivocal Positive Positive Positive Positive Positive

08-Mar 08-Mar 07-Mar 07-Mar 12-Mar 12-Mar 09-Mar 09-Mar

15-Mar 15-Mar 08-Mar 09-Mar 13-Mar 16-Mar 12-Mar 12-Mar

Molecular Molecular Serological Molecular Serological Molecular Serological Serological

B*27:09/27 group

B*27:03/05/05Q/09

B*27:05/09/27/46

Sample 1B02 was initially typed as equivocal and was further tested to determine HLA-B27+ve status using our additional test method.

Gel

B*27:03/05/05Q/09/13/17/46/49/55/56/58/60/64N/72/73/74/ 78/80/82/84/87/88/90/93/94N/96/97/98/99/104/110/111/116/ 117/118/121/122/123/124/128/132/133/135/140/141/143/14 4/146/150/151/152/155/156/158/159/162 B*27:05/13

B*27:05/09/27/46/78/125/151/154

B*27:09 Sample 1B02/2018 scored within 10 of the decision line. If this was a patient it would be referred for confirmation by PCR.

Commercial Own design

Gel Gel

Commercial

Gel

B*27:05:23, 27:18/23/29/75/85/157 are not detected by this method

Both of these samples had results in the 'grey zone'. Our laboratory procedure is to send samples to an external laboratory for testing by PCR.

Result by flow cytometer fell within the manufacturer's recommended equivocal (indeterminant) range. Commercial

Gel 1B02/2018 would be sent for high resolution HLA-B27 testing. Flow on this sample was positive but needed PCR for confirmation.

In case of a patient, sample 1B02/2018 is sent to confirmation by PCR, sample is positive. 1B01/2018 is Positive in EXON 2 and Exon 3. 1B02/2018 is Positive in Exon 2 only and Negative in Exon 3 Commercial

Gel

Commercial Commercial

Gel Gel

Commercial Commercial

Fluorescence Luminex

Commercial

Gel

Own design

Fluorescence

Commercial Own design

Gel Fluorescence

Commercial

Gel

Molecular Molecular

Commercial Commercial

Molecular Molecular Molecular Serological Molecular Molecular Molecular Serological Molecular Molecular Molecular Molecular Molecular

Commercial Commercial

Gel Coloured dots by substrate conversion on microchip Gel

Molecular Molecular Molecular Molecular Molecular Serological Molecular Serological Molecular Molecular Serological

B*27:03/05 group

By flow cytometry, the sample 1B02/2018 is negative

B*27:01-27:05:08, 27:05:10-27:05:15, 27:05:17-27:05:22, B*27:01-27:05:08, 27:05:10-27:05:15, 27:05:17-27:05:22, 27:05:24-27:05:32, 27:08/10/12-13/15-17/25/28/3627:05:24-27:05:32, 27:08/10/12-13/15-17/25/28/3640/42/44-45/47-69/71-74/79-80/82-84/86-27:90:03/9240/42/44-45/47-69/71-74/79-80/82-84/86-27:90:03/92100/103-105/108-124/126/128/131-135/137/139/141100/103-105/108-124/126/128/131-135/137/139/141149/151-152/155-156 149/151-152/155-156

Commercial

B*27:09 Commercial

Melting point Sample 1B01/2018 Positive exon 2 and exon 3 Sample 1B02/2018 only Positive exon 2

Commercial Commercial

Fluorescence Tapestation (Agilent)/Gel

Commercial

Gel

UK NEQAS for H&I is operated by Velindre NHS Trust, a UKAS proficiency testing provider No 8351. © Confidential report; no data may be published without permission from UK NEQAS for H&I

1B02/2018 was very degraded - non-interpretable result for this reason.

1B02/2018 is in the gray zone. Such results are confirmed by an alternate method. This control is positive only in exon 2

In-house HLA-B27

NEQ-115 Issue 2 P. 1 of 2 Effective Date 10/10/16

UK NEQAS FOR H&I SCHEME 1B - HLA-B27 TESTING HLA-B27 TYPING RESULTS OF SAMPLES 1B01&02/2018 DESPATCHED ON 06TH MARCH 2018 Monoclonal 1 Source of pos/neg control cells used (if applicable) Designation Specificity 20 In-house Anti-B27-FITC/CD3-PE B27+B7 UCHT1 (PE-Dyomics649) CD3 67 Anti-B27 FITC 128 GLA-ABCm3/BB7.1 HLA-B27/B7 137 HLA-B27-FITC HLA-B27-FITC 324 CD3 PE 100% 377 In-house

Lab No

Manufacturer BD Beckman Coulter Beckman Coulter Immunotech Beckman Coulter BD

Monoclonal 2 Designation Specificity Anti-B27 B27 ABCm3 and FD705 (PE) HLA-B27 Anti-B7 PE FD705 HLA-B27 HLA-B7-PE HLA-B7-PE HLA-B27 92%

Monoclonal 3 Manufacturer Designation Specificity Manufacturer One Lambda Beckman Coulter BB7.1 (unlabelled) HLA-B27 Beckman Coulter Beckman Coulter CD45 PerCP-Cy5.5 BD One Lambda Beckman Coulter BD

NEQ-115 UK NEQAS for H&I is operated by Velindre NHS Trust, a UKAS proficiency testing provider No 8351. © Confidential report; no data may be published without permission from UK NEQAS for H&I

Issue 2 P. 2 of 2 Effective Date 10/10/16